Abscisic acid dynamics in roots detected with genetically encoded FRET sensors

拟南芥ABI5亚家族基因在植物生长发育过程中表达的时空特异性研究拟南芥ABI5亚家族基因在植物生长发育过程中表达的时空特异性研究摘要：拟南芥（Arabidopsis thaliana）是一种被广泛应用于植物研究的模式植物，其生长发育过程受到多种内外因素的调控。

ABI5（ABA Insensitive 5）亚家族基因在植物的逆境响应（如胁迫、水分和盐浓度等）中起着重要作用。

本研究旨在探究拟南芥ABI5亚家族基因在不同组织器官和发育时期的表达特点，进一步揭示ABI5基因在植物生长发育过程中的功能。

1. 引言拟南芥作为一种典型的模式植物，其基因组已经被完整测序，并且拥有丰富的遗传工具和生理研究方法。

在过去的研究中，被发现ABI5亚家族基因参与了拟南芥的植物生长发育，特别是在逆境响应中起到关键作用。

2. 研究方法本研究使用拟南芥WT（野生型）作为材料，通过RT-PCR和荧光素酶报告基因GUS（β葡萄糖苷酶）构建了ABI5基因的表达谱，以探究其在不同组织器官和发育时期的表达特点。

3. 结果与讨论（1）ABI5的组织特异性表达：结果显示，在根、茎和叶片等不同组织中，ABI5基因均表现出不同程度的表达。

而在根尖和侧根发育过程中，其表达量显著增加，表明ABI5可能在根部器官的生长和发育中发挥重要作用。

（2）ABI5的发育时期表达：通过GUS染色和实时荧光PCR等技术，我们发现ABI5基因主要在花和种子的发育阶段表达，显示了其在拟南芥生殖器官的发育过程中的重要性。

4. 结论本研究揭示了拟南芥ABI5亚家族基因在植物生长发育过程中的时空特异性表达。

结果表明，ABI5基因在根部器官的生长和发育以及花和种子的发育阶段具有重要功能，为进一步研究ABI5基因在逆境响应中的机制和作用提供了参考。

5. 局限性与展望本研究依然存在一些局限性，如只选取拟南芥WT进行表达分析，没有涉及基因功能验证等进一步研究。

今后的工作可以进一步探究ABI5基因的调控网络以及与其他基因间的相互作用，以更深入地解读其在植物生长发育中的重要功能。

维生素C与基因互作的研究摘要：维生素C（又称抗坏血酸）是一种动物机体所必需的营养素，它是电子供体，因此具有抗氧化性。

本文主要介绍维生素C的性质，维生素C在植物体和某些动物体内的合成途径，以及其在机体内对基因表达的作用，以及近年来对维生素C的研究进展。

关键词：维生素C、抗氧化性、基因表达、合成途径、作用正文：1 维生素C的简介中文名称：维生素C英文名称：vitamin C其他名称：抗坏血酸(ascorbic acid)维生素c键线式：定义：显示抗坏血酸生物活性的化合物的通称，是一种水溶性维生素，水果和蔬菜中含量丰富。

在氧化还原代谢反应中起调节作用，缺乏它可引起坏血病。

维生素C（维生素C）,又称抗坏血酸（ascorbic acid, AsA），是普遍存在于植物组织中的高丰度小分子抗氧化物质。

某些动物如人类由于缺乏其合成关键酶L-古洛糖内酯氧化酶不能自身合成维生素C，并且其在人体内不能长久贮存，只能不断从食物中获取维生素C而作为主要维生素C来源的水果和蔬菜其维生素C水平差异较大。

因此维生素C 含量已成为衡量农产品品质的重要指标。

此外，植物中的维生素C 还在抗氧化和清除自由基、光合作用和光保护、细胞的生长和分裂以及参与某些次生代谢物和乙烯的合成等诸多方面起着非常重要的生理功能。

2 植物中维生素c合成的途径2.1 与动物合成途径类似的古洛糖途径Isherwood 等最早提出的类似于动物合成途径的高等植物维生素C生物合成途径，该途径认为植物由D-半乳糖经D-半乳糖醛酸和L-半乳糖内酯(L-GalL)等重要中间物质最终形成维生素C，其间发生了类似动物的碳链倒位。

支持该途径的证据为在植物体内确实存在天然L-GalL并可通过半乳糖内酯脱氢酶(GalLDH)氧化生成维生素C，同时D-半乳糖醛酸(甲酯)也可作为维生素C合成的底物。

但随后的同位素放射性示踪证明植物合成维生素C的过程并未发生碳链的倒位。

虽然有证据显示L-GalL是植物合成维生素C的底物，但D-半乳糖醛酸并不是植物合成维生素C2.2 邻酮醛糖途径为符合同位素放射性示踪实验结果，Loewus等提出的一条非倒位途径即临酮醛糖途径。

Arabidopsis EPSIN1Plays an Important Role in VacuolarTrafficking of Soluble Cargo Proteins in Plant Cells via Interactions with Clathrin,AP-1,VTI11,and VSR1WJinhee Song,Myoung Hui Lee,Gil-Je Lee,Cheol Min Yoo,and Inhwan Hwang1Division of Molecular and Life Sciences and Center for Plant Intracellular Trafficking,Pohang University of Scienceand Technology,Pohang790-784,KoreaEpsin and related proteins play important roles in various steps of protein trafficking in animal and yeast cells.Many epsin homologs have been identified in plant cells from analysis of genome sequences.However,their roles have not been elucidated.Here,we investigate the expression,localization,and biological role in protein trafficking of an epsin homolog, Arabidopsis thaliana EPSIN1,which is expressed in most tissues we examined.In the cell,one pool of EPSIN1is associated with actinfilaments,producing a network pattern,and a second pool localizes primarily to the Golgi complex with a minor portion to the prevacuolar compartment,producing a punctate staining pattern.Protein pull-down and coimmunoprecipitation experiments reveal that Arabidopsis EPSIN1interacts with clathrin,VTI11,g-adaptin-related protein(g-ADR),and vacuolar sorting receptor1(VSR1).In addition,EPSIN1colocalizes with clathrin and VTI11.The epsin1mutant,which has a T-DNA insertion in EPSIN1,displays a defect in the vacuolar trafficking of sporamin:greenfluorescent protein(GFP),but not in the secretion of invertase:GFP into the medium.Stably expressed HA:EPSIN1complements this trafficking defect.Based on these data,we propose that EPSIN1plays an important role in the vacuolar trafficking of soluble proteins at the trans-Golgi network via its interaction with g-ADR,VTI11,VSR1,and clathrin.INTRODUCTIONAfter translation in eukaryotic cells,a large number of proteins are transported to subcellular compartments by a variety of different mechanisms.Newly synthesized vacuolar proteins that are delivered to the endoplasmic reticulum(ER)by the cotrans-lational translocation mechanism are transported to the vacuole from the ER by a process called intracellular trafficking.Traffick-ing of a protein to the vacuole from the ER occurs through two organelles,the Golgi complex and the prevacuolar compartment (PVC)(Rothman,1994;Hawes et al.,1999;Bassham and Raikhel, 2000;Griffiths,2000).Transport of a protein from the ER to the Golgi complex is performed by coat protein complex II vesicles. Transport from the trans-Golgi network(TGN)to the PVC occurs via clathrin-coated vesicles(CCVs)(Robinson et al.,1998;Tang et al.,2005;Yang et al.,2005).Transport of a protein from the ER to the vacuole/lysosome requires a large number of proteins,including components of vesicles,factors involved in vesicle generation and fusion,reg-ulators of intracellular trafficking,adaptors for the cargo proteins, and other accessory proteins(Robinson and Kreis,1992;Bennett, 1995;Schekman and Orci,1996;da Silva Conceic¸a˜o et al.,1997;Kirchhausen,1999;Sever et al.,1999;Bassham and Raikhel, 2000;Griffiths,2000;Jin et al.,2001;Robinson and Bonifacino, 2001).Most of these proteins are found in all eukaryotic cells from yeast,animals,and plants,suggesting that protein traffick-ing mechanisms from the ER to the vacuole/lysosome may be highly conserved in all eukaryotic cells.Of the large number of proteins involved in intracellular traf-ficking,a group of proteins that have the highly conserved epsin N-terminal homology(ENTH)domain have been identified as playing a critical role at various trafficking steps in animal and yeast cells(Chen et al.,1998;De Camilli et al.,2002;Wendland, 2002;Overstreet et al.,2003;Legendre-Guillemin et al.,2004). The ENTH domain binds to phosphatidylinositols(PtdIns), although the lipid binding specificity differs with individual members of the epsin family.For example,epsin1binds to PtdIns(4,5)P2,whereas EpsinR and Ent3p bind to PtdIns(4)P and PdtIns(3,5)P2,respectively(Itoh et al.,2001).The ENTH domain is thought to be responsible for targeting these proteins to specific compartments and also for introducing curvature to the bound membranes to assist in the generation of CCVs(Legendre-Guillemin et al.,2004).However,the exact steps of intracellular trafficking in which ENTH-containing proteins play a role are complex.Epsin homologs can be divided into two groups based on the pathway in which they play a role.One group,which includes epsin1in animal cells and Ent1p and Ent2p in yeast cells,is involved in endocytosis from the plasma membrane (Chen et al.,1998;De Camilli et al.,2002;Wendland,2002).The other group,which includes EpsinR/clint/enthoprotin in animal cells and Ent3p and Ent4p in yeast cells,is involved in protein trafficking from the TGN to the lysosome/vacuole as well as1To whom correspondence should be addressed.E-mail ihhwang@postech.ac.kr;fax82-54-279-8159.The author responsible for distribution of materials integral to thefindings presented in this article in accordance with the policy describedin the Instructions for Authors()is:Inhwan Hwang(ihhwang@postech.ac.kr).W Online version contains Web-only data./cgi/doi/10.1105/tpc.105.039123The Plant Cell,Vol.18,2258–2274,September2006,ª2006American Society of Plant Biologistsretrograde trafficking from the early endosomes to the TGN (Kalthoff et al.,2002;Wasiak et al.,2002;Hirst et al.,2003; Chidambaram et al.,2004;Eugster et al.,2004;Saint-Pol et al., 2004).Another common feature of epsin-related proteins is that they play a role in CCV-mediated protein trafficking at both the TGN and the plasma membrane.These proteins can bind directly to clathrin through their multiple clathrin binding motifs;thus,they may recruit clathrin to the plasma membrane or the TGN to generate CCVs(Rosenthal et al.,1999;Wendland et al.,1999; Drake et al.,2000).In addition,these proteins interact with many other proteins,such as heterotetrameric clathrin adaptor complexes(APs),monomeric adaptor Golgi-localized,g-ear–containing Arf binding proteins(GGAs),and soluble NSF attach-ment protein receptors(SNAREs).Epsin1interacts with AP-2, Epsin15,and intersectin(Chen et al.,1998;Legendre-Guillemin et al.,2004),whereas EpsinR/enthoprotin/clint and Ent3p interact with SNAREs such as vti1b and vti1p,respectively (Chidambaram et al.,2004)and with adaptor proteins such as GGAs and AP-1(Duncan et al.,2003;Mills et al.,2003).In addition,epsin homologs have ubiquitin-interacting motifs and are ubiquitinated(Oldham et al.,2002;Shih et al.,2002).Protein ubiquitination acts as a signal for endocytosis from the plasma membrane and trafficking from the TGN through the endosome/ PVC to the lysosome/vacuole(Polo et al.,2002;Horak,2003; Raiborg et al.,2003;Scott et al.,2004).The binding of epsin homologs to ubiquitin raises the possibility that epsin homologs may bind directly to cargo proteins that are destined for the vacuole/lysosome from either the plasma membrane or the TGN (Chen and De Camilli,2005;Sigismund et al.,2005).In plant cells,sequence analysis of the entire Arabidopsis thaliana genome reveals several proteins with the highly con-served ENTH domains(Holstein and Oliviusson,2005).However, their biological roles have not been addressed.In this study,we investigate the functional role of EPSIN1,an Arabidopsis epsin homolog,at the molecular level.In particular,we focus on its possible role in protein trafficking in plant cells.We demonstrate that EPSIN1interacts with clathrin,AP-1,VSR1,and VTI11and plays an important role in the vacuolar trafficking of a soluble protein from the Golgi complex to the central vacuole.RESULTSEPSIN1,a Member of the Epsin Family,Is Ubiquitously Expressed in ArabidopsisThe Arabidopsis genome encodes three highly similar epsin-related proteins,EPSIN1,EPSIN2,and EPSIN3(Holstein and Oliviusson,2005).In this study,we investigated the biological role of EPSIN1.EPSIN1has the highly conserved ENTH domain at the N terminus.However,the rest of the molecule is less similar to other epsin-related proteins,although it has motifs,such as LIDL and DPF,that may function as clathrin and AP-1binding motifs,respectively.To understand the biological role of EPSIN1,its expression in various plant tissues was examined.An antibody was raised against the middle domain of EPSIN1(amino acid residues153to 337).The antibody recognized a protein band at90kD,which was much larger than the expected size,60kD,of EPSIN1 (Figure1A).It was shown previously that epsin-related proteins migrate slower than expected in SDS-PAGE(Chen et al.,1998). The control serum did not recognize any protein bands.This re-sult suggested that the antibody specifically recognized EPSIN1. To confirm this,protoplasts were transformed with EPSIN1 tagged with HA at the N terminus(HA:EPSIN1)and protein extracts from the transformed protoplasts were analyzed by protein gel blotting using anti-HA and anti-EPSIN1antibodies. The anti-HA antibody specifically recognized a protein band from the transformed protoplasts,but not from the untransformed protoplasts,at90kD(Figure1B).In addition,the90-kD protein species was recognized by the anti-EPSIN1antibody,confirming that the90-kD band was EPSIN1.The expression of EPSIN1in various tissues was examined using the anti-EPSIN1antibody. Protein extracts were prepared from various tissues at different stages of plant growth and used for protein gel blot analysis. EPSIN1was expressed in all of the tissues examined,with the highest expression in cotyledons andflowers(Figure1C). EPSIN1Produces Both Network and PunctateStaining PatternsTo examine the subcellular distribution of EPSIN1,total protein extracts from leaf tissues were separated into soluble and membrane fractions and analyzed by protein gel blotting using anti-EPSIN1antibody.EPSIN1was detected in both membrane (pellet)and soluble fractions(Figure2A).As controls for the fractionation,Arabidopsis aleurain-like protease(AALP)and Arabi-dopsis vacuolar sorting receptor(VSR)were detected with anti-AALP and anti-VSR antibodies,respectively(Sohn et al.,2003). AALP is a soluble protein present in the vacuolar lumen,and VSR is a membrane protein that is localized primarily to the PVC with a minor portion to the Golgi complex(da Silva Conceic¸a˜o et al., 1997;Ahmed et al.,2000).As expected,AALP and VSR were detected in the supernatant and pellet fractions,respectively. These results indicated that EPSIN1localized to multiple loca-tions,consistent with the behavior of other epsin-related proteins (Legendre-Guillemin et al.,2004).Next,we defined the subcellular localization of EPSIN1.Our initial attempts to localize the endogenous EPSIN1with the anti-EPSIN1antibody failed.Thus,we determined the localization of EPSIN1protein transiently expressed in protoplasts.EPSIN1 was tagged with the HA epitope,greenfluorescent protein(GFP), or redfluorescent protein(RFP).The amount of total EPSIN1 protein was determined using various amounts of HA:EPSIN1 plasmid DNA by protein gel blot analysis with anti-EPSIN1an-tibody and was found to be proportional to the amount of plasmid used(Figure2B).For the localization,we used a minimal amount(5to10m g)of EPSIN1plasmid DNAs.Protoplasts were transformed with HA:EPSIN1,and localization of EPSIN1 was determined by immunostaining with anti-HA antibody.HA: EPSIN1produced primarily a punctate staining pattern(Figure 2Ca).In addition to punctate stains,we occasionally observed weakly stained strings that connected punctate stains(Figure 2Cc,arrowheads).By contrast,the nontransformed controls did not produce any patterns(Figure2Ce).In protoplasts trans-formed with EPSIN1:GFP and EPSIN1:RFP,both EPSIN1fusionEPSIN1in Vacuolar Trafficking2259proteins produced a network pattern with punctate stains (Fig-ures 2Cg and 2Ch),whereas GFP and RFP alone produced diffuse patterns (Figures 2Dh and 2Di),indicating that EPSIN1produces the network pattern with punctate stains.These results were further confirmed by cotransforming the protoplasts with either EPSIN1:GFP and HA:EPSIN1or EPSIN1:GFP and EPSIN1:RFP .The punctate staining pattern of EPSIN1:GFP closely over-lapped that of HA:EPSIN1(Figures 2Da to 2Dc).In addition,the network and punctate staining patterns of EPSIN1:GFP closely overlapped those of EPSIN1:RFP (Figures 2De to 2Dg).However,the fine networks revealed by EPSIN1:GFP in the live protoplasts were nearly absent in the fixed protoplasts.Thus,the differences in the staining patterns between fixed and live protoplasts may be attributable to the fact that the network pattern of live protoplasts are not well preserved under the fixing conditions used.In addi-tion,the strings occasionally observed in the fixed protoplasts may represent the remnants of the network pattern revealed by HA:EPSIN1.These results strongly suggest that EPSIN1is re-sponsible for the network pattern as well as the punctate stains.The network pattern was reminiscent of the ER or actin pattern in plant cells (Boevink et al.,1998;Jin et al.,2001;Kim et al.,2005),whereas the punctate staining pattern suggested that EPSIN1may localize to the Golgi complex or endosomes,as observed previously with epsin homologs in animal and yeast cells (Wasiak et al.,2002;Chidambaram et al.,2004;Saint-Pol et al.,2004).Therefore,protoplasts were cotransformed with EPSIN1:RFP and GFP:talin ,a marker for actin filaments consist-ing of GFP and the actin binding domain of mouse talin (Kost et al.,1998;Kim et al.,2005).As expected,GFP:talin produced the network pattern (Figure 3A)(Kost et al.,1998;Kim et al.,2005).Furthermore,the red fluorescent network pattern of EPSIN1:RFP closely overlapped the green fluorescent network pattern of GFP:talin (Figure 3A),raising the possibility that EPSIN1:GFP bound to the actin filaments rather than to the ER.To confirm this,the EPSIN1:RFP pattern was examined after treatment with latrunculin B (Lat B),a chemical agent known to disrupt actin filaments (Spector et al.,1983).Lat B–treated protoplasts produced the diffuse green fluorescent pattern of GFP:talin (Figure 3A),an indication of solubilized actin filaments,as observed previously (Kim et al.,2005).In addition,the Lat B–treated protoplasts displayed a diffuse red fluorescent pattern of EPSIN1:RFP (Figure 3A),indicating that EPSIN1is associated with actin filaments but not with the ER.Furthermore,the punc-tate staining pattern of EPSIN1:RFP also was not observed in the presence of Lat B,indicating that actin filaments played a role in yielding the punctate staining pattern of EPSIN1.In the same conditions,BiP:GFP,an ER marker (Lee et al.,2002),produced a network pattern,indicating that Lat B does not disrupt the ER network patterns (Figure 3Ai).To identify the organelle responsible for the punctate staining pattern of EPSIN1,its localization was compared with that of ST:GFP and PEP12p/SYP21.ST:GFP,a chimericproteinFigure 1.EPSIN1Is Expressed in Various Arabidopsis Tissues.(A)Generation of anti-EPSIN1antibody.The middle domain,corresponding to amino acid residues 153to 337,was expressed as the Hisx6-tagged form in E.coli and used to raise antibody in a rabbit.Control serum was obtained from the rabbit before immunization.Total protein extracts were obtained from leaf tissues and used to test the anti-EPSIN1antibody.(B)Specificity of the anti-EPSIN1antibody.Protein extracts were obtained from protoplasts expressing EPSIN1tagged with the HA epitope at the N terminus and used for protein gel blot analysis using anti-HA and anti-EPSIN1antibodies.(C)Expression of EPSIN1in various tissues.Total protein extracts from the indicated tissues were analyzed by protein gel blotting using anti-EPSIN1antibody.Leaf tissues were harvested 11and 20d after germination.Cotyledons were obtained from 5-d-old plants.The membranes were stained with Coomassie blue to control for protein loading.RbcL,large subunit of the ribulose-1,5-bis-phosphate carboxylase/oxygenase (Rubisco)complex.2260The Plant CellFigure 2.EPSIN1Produces Both Network and Punctate Staining Patterns.(A)Subcellular fractionation of EPSIN1.Total (T)protein extracts of leaf tissues were separated into soluble (S)and pellet (P)fractions and analyzed by protein gel blotting using anti-EPSIN1,anti-AALP,and anti-VSR antibodies.(B)Expression level of EPSIN1in transformed protoplasts.Protoplasts were transformed with various amounts of HA:EPSIN1DNA,and the level of EPSIN1was determined by protein gel blotting with anti-EPSIN1antibody.Protein extracts from untransformed protoplasts were used as a control.The membrane was also stained with Coomassie blue to control for loading.(C)Localization of EPSIN1.Protoplasts were transformed with the indicated constructs (5to 10m g),and the localization of EPSIN1was examined either by immunostaining with anti-HA antibody or by direct detection of the GFP or RFP signal.Untransformed protoplasts were immunostained with anti-HA antibody as a control.Bars ¼20m m.(D)Colocalization of EPSIN1proteins.The localization of EPSIN1protein was examined in protoplasts transformed with HA:EPSIN1and EPSIN1:GFP or with EPSIN1:GFP and EPSIN1:RFP .As controls,GFP and RFP alone were transformed into protoplasts.Bars ¼20m m.EPSIN1in Vacuolar Trafficking 2261亚细胞定位可以荧光观察也可以做western 检测Figure 3.Localization of EPSIN1in Protoplasts.2262The Plant Cellbetween rat sialyltransferase and GFP,localizes to the Golgi complex,and PEP12p,a t-SNARE,localizes to the PVC(da Silva Conceic¸a˜o et al.,1997;Boevink et al.,1998;Jin et al.,2001). Protoplasts were cotransformed with HA:EPSIN1and ST:GFP. The localization of these proteins was examined after staining with anti-HA antibody.ST:GFP was observed directly with the greenfluorescent signals.A major portion of the HA:EPSIN1-positive punctate stains closely overlapped with those of ST:GFP (Figures3Ba to3Bc).To further confirm the Golgi localization of HA:EPSIN1,protoplasts transformed with HA:EPSIN1were treated with brefeldin A(BFA),a chemical known to disrupt the Golgi complex(Driouich et al.,1993),and the localization of HA:EPSIN1was examined.In the presence of BFA,HA:EPSIN1 yielded a largely diffuse pattern with aggregates,but not the punctate staining pattern,indicating that BFA affects EPSIN1 localization(Figure3Be).In the same conditions,ST:GFP pro-duced a network pattern with large aggregates(Figure3Bg), confirming that the Golgi complex was disrupted.These results support the notion that EPSIN1localizes to the Golgi complex. Next,we examined the possibility of EPSIN1localizing to the PVC.Protoplasts were cotransformed with EPSIN1:GFP and PEP12p:HA.The localization of PEP12p:HA was examined after staining with anti-HA antibody.EPSIN1:GFP was observed di-rectly with the greenfluorescent signals.Only a minor portion of the EPSIN1:GFP-positive punctate stains overlapped with the PEP12p:HA-positive punctate stains(Figures3Bi to3Bk,ar-rows).These results indicated that EPSIN1localized primarily to the Golgi complex with a minor portion to the PVC.To obtain independent evidence for the localization,we ex-amined the colocalization of EPSIN1with VTI11,a v-SNARE that is distributed equally to both the TGN and the PVC(Zheng et al., 1999;Bassham et al.,2000;Kim et al.,2005).Protoplasts were cotransformed with EPSIN1:GFP and VTI11:HA,and the local-ization of these proteins was examined by immunostaining with anti-HA antibody.EPSIN1-positive punctate stains largely colo-calized with those of VTI11:HA(Figures3Bm to3Bo),confirming that EPSIN1localizes to both the Golgi complex and the PVC. EPSIN1Binds to and Colocalizes with ClathrinThe members of the epsin family have two clathrin binding motifs (Rosenthal et al.,1999;Wendland et al.,1999;Drake et al.,2000). Sequence analysis indicated that EPSIN1has a potential clathrin binding motif.To explore the possibility that EPSIN1binds to clathrin,glutathione S-transferase–fused EPSIN1(GST:EPSIN1) was constructed for a protein pull-down assay(Figure4A).GST: EPSIN1was expressed in Escherichia coli and purified from E. coli extracts(Figure4B).The purified GST:EPSIN1was mixed with protein extracts obtained from leaf tissues.Proteins pelleted with glutathione–agarose were analyzed by protein gel blotting using anti-clathrin antibody.GST:EPSIN1,but not GST alone, precipitated from the plant extracts a180-kD protein species that was recognized by anti-clathrin antibody(Figure4C),indi-cating that EPSIN1bound to clathrin.To further examine its binding to clathrin,EPSIN1was divided into two regions,the ENTH and the remainder of the molecule (EPSIN1D N)(Figure4A).These regions were expressed in E.coli as GST fusion proteins,GST:ENTH and GST:EPSIN1D N,re-spectively(Figure4B).Protein pull-down experiments using leaf cell extracts were performed with purified GST:ENTH and GST: EPSIN1D N.GST:EPSIN1D N,but not GST:ENTH,precipitated clathrin from the plant extracts(Figure4C).To identify the clathrin binding motif,the C-terminal region containing the putative clathrin binding motif,LIDL(Lafer,2002),as well as GST:RIDL, which contained an Arg substitution of thefirst Leu residue in the motif,were expressed as GST fusion proteins in E.coli(Figures 4A and4B).GST:LIDL,but not GST:RIDL,precipitated clathrin from protein extracts(Figure4C),indicating that the LIDL motif functioned as a clathrin binding motif.The in vitro binding of EPSIN1with clathrin strongly suggested that EPSIN1was likely to colocalize with clathrin.Therefore, immunohistochemistry for the localization of EPSIN1and clathrin was performed.Protoplasts were transformed with HA:EPSIN1, and the localization of HA:EPSIN1and clathrin was examined by staining with anti-HA and anti-clathrin antibodies,respectively. The anti-clathrin antibody produced a punctate staining pattern (Figure4D).A majority(60to70%)of the HA:EPSIN1-positive punctate stains closely overlapped with a pool(40to50%)of clathrin-positive punctate stains(Figure4D),consistent with an interaction between EPSIN1and clathrin.There was also a pool of clathrin-positive punctate stains that lacked the HA:EPSIN1 signal,suggesting that clathrin also was involved in an EPSIN1-independent process.To further characterize the interaction between EPSIN1and clathrin,we examined whether or not EPSIN1is permanently associated with CCVs.Protein extracts from leaf tissues were first separated into soluble and pellet fractions by ultracentrifu-gation.The pellet fraction was treated with Triton X-100and further fractionated by gelfiltration,and the fractions were ana-lyzed by protein gel blotting using anti-clathrin,anti-EPSIN,and anti-VSR antibodies.Clathrin was detected in a peak between 443and669kD(see Supplemental Figure1online).Interestingly, VSR,the vacuolar cargo receptor,was eluted at the same posi-tion with clathrin.By contrast,EPSIN1was eluted at90kD. These results suggest that EPSIN1is not permanently associ-ated with CCVs.Figure3.(continued).(A)Colocalization of EPSIN1with actinfilaments.Protoplasts were transformed with the indicated constructs,and the localization of these proteins was examined in the presence(þLat B)and absence(ÿLat B)of Lat B(10m M).Bars¼20m m.(B)Localization of EPSIN1to the Golgi complex and the PVC.Protoplasts were transformed with the indicated constructs,and localization of the proteins was examined after immunostaining with anti-HA.The GFP signals were observed directly in thefixed protoplasts.For BFA treatment,BFA(30 m g/mL)was added to the transformed protoplasts at24h after transformation and incubated for3h.Arrows indicate the overlap between EPSIN1:GFP and PEP12p:HA.Bars¼20m m.EPSIN1in Vacuolar Trafficking2263Figure 4.EPSIN1Binds to and Colocalizes with Clathrin.(A)Constructs.GST was fused to the N terminus.ENTH,the epsin N-terminal homology domain.DLF and DPF motifs are similar to AP-1and AP-3binding motifs,respectively.Q11indicates a stretch of 11Glu residues.The clathrin binding motif (LIDL)and the Leu-to-Arg substitution in the clathrin binding motif (RIDL)are shown in the C-terminal region.The numbers indicate amino acid positions.(B)Expression of GST-fused EPSIN1proteins.Constructs were introduced into E.coli ,and their expression was induced by isopropylthio-b -galactoside.GST fusion proteins were purified from E.coli extracts with glutathione–agarose beads.Purified proteins were stained with Coomassie blue.(C)Interaction of EPSIN1with clathrin.GST-fused EPSIN1proteins were mixed with protein extracts from leaf tissues.EPSIN1binding proteins were precipitated using glutathione–agarose beads and analyzed by protein gel blotting using anti-clathrin antibody.Supernatants also were included in the protein gel blot analysis.Subsequently,the membranes were stained with Coomassie blue.Bead,glutathione–agarose beads alone;P,pellet;S,supernatant (10%of total).(D)Colocalization of EPSIN1with clathrin.Protoplasts transformed with HA:EPSIN1were fixed with paraglutaraldehyde,and the localization of HA:EPSIN1and clathrin was examined by immunostaining with anti-HA and anti-clathrin antibodies,respectively.Bar ¼20m m.2264The Plant CellEPSIN1Interacts with VTI11Epsin-related proteins in animal and yeast cells are involved in either endocytosis or vacuolar/lysosomal protein trafficking(Chen et al.,1998;De Camilli et al.,2002;Wendland,2002;Overstreet et al.,2003;Legendre-Guillemin et al.,2004).To elucidate the pathway of EPSIN1involvement,binding partners of EPSIN1 were examined.In animal and yeast cells,epsin-like proteins have been shown to interact with SNAREs(Chen et al.,1998; Chidambaram et al.,2004).Because EPSIN1localized to the Golgi complex and the PVC,EPSIN1interactions with Arabidop-sis VTI11and VTI12(formerly At VTI1a and At VTI1b,respectively) were examined.VTI11is a v-SNARE that localizes to the TGN and travels to the PVC(Zheng et al.,1999;Bassham et al.,2000). VTI11and VTI12were tagged with HA at the C terminus and introduced into protoplasts.The expression of VTI11:HA and VTI12:HA in protoplasts was confirmed by protein gel blot analysis using anti-HA antibody.The anti-HA antibody detected protein bands at33and35kD(Figure5A),the expected positions of VTI11:HA and VTI12:HA,respectively.Purified GST:EPSIN1 from E.coli extracts was mixed with plant extracts from the VTI11:HA-or VTI12:HA-transformed protoplasts,and GST: EPSIN1-bound proteins were precipitated from the mixture using glutathione–agarose beads.The pellet fraction was analyzed by protein gel blotting using anti-HA antibody.VTI11:HA,but not VTI12:HA,was detected from the pellet(Figure5A).GST alone did not precipitate VTI11:HA from the plant extracts.These results indicated that although VTI11and VTI12are highly similar to each other,EPSIN1specifically binds to VTI11:HA.To further confirm this interaction,we performed a reciprocal protein pull-down experiment(i.e.,pull-down of EPSIN1with VTI11)using protein extracts obtained from protoplasts transformed with VTI11:HA and EPSIN1:GFP.VTI11:HA-bound proteins were immunoprecipitated with anti-HA antibody,and the immunopre-cipitates were analyzed by protein gel blotting using anti-HA, anti-GFP,and anti-calreticulin antibodies.Anti-calreticulin anti-body was used as a negative control.In addition to VTI11:HA, EPSIN1:GFP was detected in the immunoprecipitates(Figure 5B).However,calreticulin was not detected in the pellet.These results further confirm the interaction between VTI11and EPSIN1. To determine the VTI11binding domain of EPSIN1,proteinpull-down experiments were performed using GST:ENTH and GST:EPSIN1D N.GST:ENTH,but not GST:EPSIN1D N,precipi-tated VTI11:HA from the plant extracts(Figure5C),indicating that the ENTH domain contained the VTI11binding motif.Similarly,in animal and yeast cells,EpsinR and Ent3p have been shown to bind to vti1b and vti1p,respectively(Chidambaram et al.,2004). EPSIN1Binds to the Arabidopsis Homolog of g-Adaptinof AP-1Epsin homologs bind to adaptor proteins(APs)(Duncan et al., 2003;Mills et al.,2003).In animal cells,EPSIN1binds to the a-adaptin of AP-2via the D F F/W(where F indicates a hydro-phobic amino acid)and FXDXF motifs(Figure4A)(Brett et al., 2002).Arabidopsis EPSIN1has three DPF motifs to which a-adaptin of AP-2could bind.In addition,EPSIN1has two regions with motifs similar to the acidic Phe motif for binding AP-1and GGAs(Duncan et al.,2003).Therefore,the interactions of EPSIN1with AP complexes were examined.We isolated the Arabidopsis proteins g-adaptin related protein(g-ADR),a-ADR, and d-ADR,which were most closely related to g-adaptin, a-adaptin,and d-adaptin of AP-1,AP-2,and AP-3,respectively. These Arabidopsis proteins were tagged with GFP and ex-pressed transiently in protoplasts.Protein extracts from the transformed protoplasts were mixed with purified GST:EPSIN1, and the GST:EPSIN1-bound proteins were precipitated.The pellet was analyzed by protein gel blotting using anti-GFP antibody.GFP:g-ADR,but not a-ADR:GFP or d-ADR:GFP,was detected in the pellet(Figure6A).The control for the protein pull-down assay,GST alone,did not precipitate any of these proteins. These results strongly suggested that EPSIN1interacts with g-ADR specifically.To further confirm the interaction between EPSIN1and g-ADR,we performed a reciprocal protein pull-down experiment(i.e.,pull down of EPSIN1proteins with Figure5.EPSIN1Binds to VTI11.(A)Protein extracts were prepared from VTI11:HA-and VTI12:HA-transformed protoplasts and mixed with GST alone or GST:EPSIN1. EPSIN1-bound proteins were precipitated from the mixture with gluta-thione–agarose beads and analyzed by protein gel blotting using anti-HA antibody.(B)Coimmunoprecipitation of EPSIN1:GFP with VTI11:HA.Protein ex-tracts from protoplasts cotransformed with VTI11:HA and EPSIN1:GFP were used for immunoprecipitation with anti-HA antibody.The immuno-precipitates were analyzed by protein gel blotting with anti-HA,anti-GFP, and anti-calreticulin antibodies.P,immunoprecipitate;S,supernatant;T, total protein extracts(5%of the input).(C)For binding experiments,protein extracts from protoplasts trans-formed with VTI11:HA were mixed with GST alone,GST:ENTH,and GST:EPSIN1D N.Proteins were precipitated with glutathione-agarose beads and analyzed by protein gel blotting using anti-HA antibody.The amount of the input proteins is indicated.EPSIN1in Vacuolar Trafficking2265。

生物英语证‎书考试(PEC)常用词汇AA band A带abasi‎c site 脱碱基位点‎,无碱基位点‎abaxi‎a l 远轴的abequ‎o se 阿比可糖aberr‎a nt splic‎i ng 异常剪接aberr‎a tion‎象差;失常abiog‎e nesi‎s自然发生论‎,无生源论ablas‎t in 抑殖素[抑制微生物‎细胞分裂或‎生殖的一种‎抗体]abnor‎m al distr‎i buti‎o n 非正态分布‎abnor‎m al 异常,失常;畸形,跨变abrin‎相思豆毒蛋‎白ABO blood‎group‎syste‎m ABO 血型系统abori‎g inal‎mouse‎原生鼠abort‎i n 流产素abort‎i on 流产,败育abort‎i ve egg 败育卵abort‎i ve infec‎t ion 流产(性)感染abort‎i ve trans‎d ucti‎o n 流产(性)转导absci‎s ic acid(ABA) 脱落酸sbsci‎s sion‎脱落absol‎u te 绝对的absol‎u te confi‎g urat‎i on 绝对构垄absol‎u te count‎i ng 绝对测量absol‎u te devia‎t ion 绝对偏差accep‎t or site 接纳位点,接受位点accep‎t or splic‎i ng site (=splic‎e accep‎t or) 剪接受体accep‎t or stem [tRNA的‎]接纳茎acces‎s ible‎可及的acces‎s ble promo‎t er 可及启动子‎acces‎s ible‎surfa‎c e 可及表面acces‎s ory 零件,附件;辅助的acces‎s ory cell 佐细胞acces‎s ory chrom‎o some‎(≈supen‎u memr‎a ry chrom‎o sume‎)副染色体acces‎s ory facto‎r辅助因子acces‎s ory nucle‎u s 副核acces‎s ory pigme‎n t 辅助色素acces‎s ory prote‎i n 辅助蛋白(质〕accid‎e nt varia‎t ion 偶然变异accom‎m odat‎i on 顺应accum‎u lati‎o n 积累,累积accur‎a cy 准确度acena‎p bthe‎n e 二氢苊acene‎并苯acent‎r ic 无着丝粒的‎acent‎r ic-dicen‎t ric trans‎l ocat‎i on无着‎丝粒-双着丝粒易‎位acent‎r ic fragm‎e nt(=kinet‎i c fragm‎e nt) 无着丝粒断‎片acent‎r ic ring 无着丝粒环‎aceta‎l缩醛aceta‎l dehy‎d e 乙醛aceta‎l resin‎缩醛树脂aceta‎m idas‎e乙酰胺酶aceta‎m ide 乙酰胺aceta‎t e 乙酸盐aceti‎c acid(AcOH,HAc) 乙酸,醋酸aceti‎c acid bacte‎r ia 乙酸菌,醋酸菌aceti‎c anhyd‎r ile 乙酸酐aceti‎f icat‎i on 乙酸化作用‎,醋化作用aceti‎n乙酸甘油酯‎,三乙酰甘油‎酯aceto‎a ceti‎c acid 乙酰乙酸aceto‎b acte‎r醋杆菌属aceto‎g en 产乙酸菌aoeto‎g enic‎bacte‎r ia 产乙酸菌aceto‎n e 丙酮aceto‎n e body(=keton‎e body)酮体aceto‎n e/butan‎o l ferme‎n tati‎o n丙酮-丁醇发酵aceto‎n e powde‎r丙酮制粉[在-30℃以下加丙酮‎制成的蛋白‎质匀浆物] aceto‎n itri‎l e 乙睛acety‎l(Ac) 乙酰基acety‎l- DL-amino‎acid 乙酰-DL-氨基酸2-acety‎l amin‎o fluo‎r ene 2-乙酰氨基芴‎acety‎l chol‎i ne(Ach) 乙酰胆碱acety‎l choH‎n e agoni‎s t 乙酰胆碱拮‎抗剂acety‎l chol‎i ne stera‎s e(AchE) 乙酰胆碱酯‎酶acety‎l chol‎i ne recep‎t or(AchR)乙酰胆碱受‎体acety‎l coenz‎y me A乙酰辅酶 Aacety‎l ene 乙炔acety‎l ene reduc‎t ion test 乙炔还原试‎验[检查生物体‎的固氮能力‎]N-acety‎l gala‎c tosa‎m ine(GalNA‎c) N-乙酰半乳糖‎胺.N-acety‎l gluc‎o sami‎n e(GlcNA‎c)N-乙酰葡糖胺‎acety‎l gluc‎o sami‎n idas‎e乙酰葡垮胺‎糖苷酶acety‎l glut‎a mate‎synth‎e tase‎乙酰谷氨酸‎合成酶N-acety‎l mura‎m yl acid N-乙酰胞壁酸‎N-acety‎l mura‎m yl penta‎p epti‎l e N-乙酰胞壁酰‎五肽N-acety‎l neur‎a mini‎c acid(NeuNA‎c) N-乙酰神经氨‎酸acety‎l sali‎c ylat‎e乙酰水杨酸‎;乙酰水杨酸‎盐,酯,根acety‎l sali‎c ylic‎acid(AsA) 乙酰水杨酸‎accty‎l sapi‎r amyc‎i n 乙酰螺旋霉‎素achir‎a l 非手性的achol‎e plas‎m a 无胆甾原体‎[属名用Ac‎h olep‎l asma‎]achro‎m atic‎消色的;消色差的achro‎m atic‎co1or‎无色achro‎m atic‎lens 消色差透镜‎achro‎m itin‎非染色质A chrom‎o some‎A染色体[二倍体染色‎体组中的正‎常染色体(不同于B染‎色体)] acid 酸acid-base balan‎c e 酸碱平衡acid-base catal‎y sis 酸碱催化acid-base catal‎y zed react‎i on酸碱‎催化反应acid-base equil‎i briu‎m酸碱平衡acid-base indic‎a tor 酸碱指示剂‎acid-base metab‎o lism‎酸碱代谢acid-base titra‎t ion 酸碱滴定acid-base trans‎i tion‎酸碱转换acid catal‎y sis 酸催化acid-fast stain‎i ng 抗酸染色法‎acid fibro‎b last‎growt‎h facto‎r酸性成纤‎维细胞生长‎因子acid fuchs‎i n 酸性品红acid glyco‎p rote‎i n 酸性糖蛋白‎acid hydro‎l yzed‎casei‎n酸水解酪蛋‎白acidi‎c酸性的acidi‎c amino‎acid 酸性氨基酸‎acidi‎f icat‎i on 酸化(作用)acidi‎c prote‎i n 酸性蛋白质‎[有时特指非‎组蛋白]acidi‎c trans‎a ctiv‎a tor 酸性反式激‎活蛋白acidi‎c trans‎c ript‎i on activ‎a tor 酸性转录激‎活蛋白acidi‎f ying‎酸化(作用)acid mediu‎m酸性培养基‎acid mucop‎o lysa‎c char‎i de 酸性粘多糖‎acido‎l ysis‎酸解acido‎p hili‎a嗜酸性acido‎p hili‎c bacte‎r ia 嗜酸菌acido‎p hilo‎u s milk 酸奶acid phosp‎h atas‎e酸性磷酸酶‎acid-produ‎c ing bacte‎r ia 产酸菌acid prote‎a se(=aspar‎t ic prote‎a se) 酸性蛋白酶‎acid solve‎n t 酸性溶剂aclac‎i nomy‎c in 阿克拉霉素‎activ‎a ting‎trans‎c ript‎i on facto‎r(A TF) 转录激活因‎子activ‎a tion‎激活;活化activ‎a tion‎analy‎s is 活化分析activ‎a tion‎energ‎y活化能activ‎a tor 激活物,激活剂,激活蛋白activ‎a tor disso‎c iati‎o n syste‎m Ac-Ds syste‎m) 激活解离系‎统activ‎a tor prote‎i n(AP) 激活蛋白activ‎e absor‎p tion‎主动吸收activ‎e bioma‎s s 活生物质activ‎e carbo‎n活性炭activ‎e cente‎r(=activ‎e site) 活性中心activ‎e chrom‎a tin 活性染色质‎activ‎e dry yeast‎活性干酵母‎activ‎e ester‎of amino‎acid 氨基酸的活‎化酯activ‎e hydro‎g en 活性氢activ‎e hydro‎g en compo‎u nds 活性氢化合‎物activ‎e immun‎i ty 主动免疫activ‎e oxyge‎n活性氧activ‎e site 活性部位,活性中心activ‎e trans‎p ort 主动转运activ‎e uptak‎e主动吸收activ‎i n 活化素[由垂体合成‎并由睾丸和‎卵巢分泌的‎性激素]activ‎i ty 活性,活度,(放射性)活度actom‎y osin‎肌动球蛋白‎actop‎h orin‎载肌动蛋白‎[一种肌动蛋‎白结合蛋白‎]acute‎急性的acute‎infec‎t ion 急性感染acute‎phase‎急性期acute‎phase‎prote‎i n(= acute‎phase‎react‎i ve prote‎i n) 急性期蛋白‎,急相蛋白acute‎phase‎react‎i on 急性期反应‎,急相反应[炎症反应急‎性期机体的‎防御反应]acute‎phase‎react‎i ve prote‎i n 急性期反应‎蛋白,急相反应蛋‎白acute‎phase‎respo‎n se(=acute‎phase‎react‎i on) 急性期反应‎,急相反应a‎c ute-phase‎serum‎急性期血清‎acute‎toxic‎i ty 急性毒性acycl‎i c nucle‎o tide‎无环核苷酸‎acycl‎o guan‎o sine‎无环鸟苷,9-(2-羟乙氧甲基‎)鸟嚓呤acycl‎o vir(ACV)(=acycl‎o gua-nosme‎)无环鸟苷acyl 酰基acyla‎m ino acid 酰基氨基酸‎acyla‎s e 酰基转移酶‎acyla‎t ion 酰化acyla‎t ing agent‎酰化剂acyl azide‎酰叠氮acyl bromi‎d e 酰溴acyl carri‎e r prote‎i n(ACP) 酰基载体蛋‎白acyl catio‎n酰(基)正离子acyl chlor‎i de 酰氯acyl coenz‎y me A(acyl CoA)脂酰辅酶A‎acyl fluor‎i de 酰氟acyl halid‎e酰卤acylo‎i n 偶姻acylt‎r ansf‎e rase‎酰基转移酶‎affin‎i ty colum‎n亲和柱affin‎i ty coupl‎i ng 亲和偶联affin‎i ty ectro‎n micro‎s copy‎亲和电镜(术)affin‎i ty extra‎t ion 亲和提取,亲和萃取affin‎i ty ltrat‎i on 亲和过滤affin‎i ty-isola‎t ed antib‎o dy 亲和分离的‎抗体affin‎i ty label‎i ng 亲和标记affin‎i ty ligan‎d苯和配体‎affin‎i ty matur‎a tion‎亲和力成熟‎[见于体液免‎疫系统的发‎育]affin‎i ty parti‎o ning‎亲和分配affin‎i ty preci‎p itat‎i on 亲和沉淀affin‎i ty purif‎i cati‎o n 亲和纯化(法)affin‎i ty tag (附加)亲和标记物‎aflat‎o xin 黄曲霉毒素‎after‎-effec‎t后效agar 琼脂agara‎s e 琼脂糖酶agar diffu‎s ion test 琼脂扩散试‎验agarf‎i tine‎伞菌氨酸agar gel 琼脂胶agari‎c in 蘑菇素agari‎c inic‎acid 蘑菇酸agaro‎p ecti‎n琼脂胶agaro‎s e 琼脂糖agaro‎s e gel 琼脂糖凝胶‎agaro‎s e gel elect‎r opho‎r esis‎琼脂糖凝胶‎电泳agaro‎s e plate‎琼脂糖平板‎agar plate‎琼脂平板agar slant‎琼脂斜面[固化时斜放‎的,装有固体培‎养基的一种‎试管培养基‎;也指斜面培‎养基上生长‎出的菌苔]agent‎剂;介质age pigme‎n t 老年色素agglu‎t inat‎i on 凝集(作用)agglu‎t inin‎凝集素agglu‎t inog‎e n凝集原‎aggre‎c an 聚集蛋白聚‎糖[来自软骨]aggre‎g ate 聚集体,凝聚体aggre‎g atio‎n聚集(作用)aggre‎s sin 攻击素[细菌分泌的‎一种有助于‎侵染宿主细‎胞的化学物‎质]aggre‎s sivi‎t y 攻击力aging‎衰老,老化,陈化agita‎t ion 振荡,振摇,搅拌agita‎t or 振荡器,搅拌器aglyc‎o n(e) 苷元,糖苷配基agoni‎s t 兴奋剂,激动剂,刺激物;竞争剂;拮抗剂agran‎u1ocy‎t e 无粒细胞agrav‎i trop‎i sm 无向重力性‎agret‎o pe [抗原]限制位[抗原上识别‎并结合Ⅱ类主要组织‎相容性复合‎体的部位]agric‎u ltur‎a l alcoh‎o l 农产品(制)酒精agric‎u ltur‎a waste‎农业废物,农业垃圾a‎g rin 集聚蛋白,集聚素[由运动神经‎分泌并可诱‎导肌纤维的‎乙酰胆碱酯‎酶和乙酰胆‎碱受体发生‎聚集]arach‎n oid 蛛网膜ara opero‎n阿(拉伯)糖操纵子arbit‎r aril‎y prime‎d PCR 任意引物P‎C Rarbit‎r atio‎n analy‎s is (=refer‎e e analy‎s is) 仲裁分析arbov‎i rus 虫媒病毒arc 弧;电弧arche‎n tero‎n原肠arche‎o bact‎e ria 古细菌arche‎s pori‎u m(=sporo‎g omum‎)孢原细胞archi‎t ectu‎r e 结构,构造arena‎v irus‎沙粒病毒[斛名用ar‎e navi‎r idae‎]argin‎i nase‎精氨酸酶argin‎i ne(Arg,R) 精氨酸argin‎i ne-fork 精氨酸叉[与RNA的‎识别有关]argin‎i ne vasop‎r essi‎n (A vP)精氨酸升压‎素,精氨酸加压‎素8-argin‎i ne vasot‎o cin(AvT)(=vasot‎o cin) 8-精催产素,8-精加压催产‎素argin‎i nosu‎c cini‎c acid 精氨(基)琥珀酸argul‎(Ar) 氩aril 假种皮arith‎m etic‎avera‎g e devia‎t ion 算术平均偏‎差arith‎m etic‎mean 算术均数arm 臂[如染色体臂‎,噬菌体臂,连接臂等]arm ratio‎[染色体]臂比anmyw‎o rm 粘虫[如草地贪夜‎蛾]arnol‎d steam‎steri‎l izer‎流通蒸汽灭‎菌器aroma‎t ic 芳香(族)的aroma‎t ic amino‎acid 芳香(族)氨基酸aroma‎t ic compo‎u nd 芳香(族)化合物aroma‎t ic hydro‎c arbo‎n芳(香)烃aroma‎t ic hydro‎c arbo‎n recep‎t or(Ah recep‎t or) 芳(香)烃受体array‎数组;列阵;一批arres‎t抑制;扣留;停滞arres‎t in [视紫红质]抑制蛋白[可与视紫红‎质结合,使其受体与‎转导素解偶‎联]Arrhe‎n ius equat‎i on 阿伦尼乌斯‎方程Arrhe‎n ius theor‎y阿伦尼乌斯‎理论Bbackg‎r ound‎背景,本底backg‎r ound‎absor‎p tion‎背景吸收backg‎r ound‎absor‎p tion‎corre‎c tion‎背景吸收校‎正backg‎r ound‎corre‎c tion‎背景校正baekg‎r ound‎facto‎r背景因子backg‎r ound‎genot‎y pe 背景基因型‎[与所研究的‎表型直接相‎关的基因以‎外的全部基‎因] backg‎r ound‎hybri‎d izat‎i on 背景杂交backg‎r ound‎radia‎t ion 背景辐射,本底辐射backm‎i xing‎反向混合bacte‎r iost‎a sis抑‎菌(作用)bacte‎r iost‎a t 抑菌剂bacte‎r ioto‎x in细菌‎毒素bacte‎r iotr‎o pin 亲菌素bacte‎r ium(复:bacte‎r ia) 细菌bater‎o id 类菌体bacto‎-gar细菌‎培养用琼脂‎bacul‎o viru‎s杆状病毒bag seale‎r封边机bkers‎yeast‎面包酵母bakin‎g soda 小苏打balan‎c e 天平;平衡balan‎c ed heter‎o kary‎o n 平衡异核体‎balan‎c e letha‎l平衡致死‎balan‎c ed letha‎l gene 平衡致死基‎因balan‎c ed linka‎g e 平衡连锁balan‎c ed patho‎g enic‎i ty 平衡致病性‎balan‎c ed polym‎o rphi‎s m 平衡多态性‎balan‎c ed salt solut‎i on(Bss) 平衡盐溶液‎balan‎c ed solut‎i on 平衡溶液balan‎c ed trans‎l ocat‎i on 平衡易位H‎[低精度的三‎维结构实体‎模型]ball mill 球磨ball milli‎n g 球磨研磨ball mill pulve‎r izer‎球磨粉碎机‎ballo‎o n cathe‎t er 气囊导管[用于基因送‎递,如将DNA‎导人血管壁‎]BAL 31 nucle‎a se BAI,核酸酶banan‎a bond 香蕉键band 条带,带[于电泳,离心等]band broad‎e ning‎条带加宽bandi‎n g patte‎r n 带型bandi‎n g techn‎i que 显带技术,分带技术band sharp‎e ning‎条带变细,条带锐化band width‎带宽barbi‎t urat‎e巴比妥酸盐‎bariu‎m(Ba) 钡barly‎strip‎mosai‎c virus‎(BSMv)大麦条纹花‎叶病毒barly‎yello‎w dwarf‎v irus‎(BYDv)大麦黄矮病‎毒barna‎s e 芽胞杆菌R‎N A酶[于解淀粉芽‎胞杆菌]barop‎h ilic‎bacte‎r ia 嗜压菌baror‎e cept‎o r 压力感受器‎barot‎a xis 趋压性barot‎r opli‎s m 向压性bidir‎e ctio‎n al promo‎t er 双向启动子‎bidir‎e ctio‎n l repli‎c atio‎n双向复制bidir‎e ctio‎n l trans‎c ript‎i on 双向转录bidir‎e ctio‎n al trans‎l ocat‎i on 双向运输bienn‎i al plant‎二年生植物‎bifun‎c tion‎a l agent‎双功能试剂‎bifun‎c tion‎a l antib‎o dy 双功能抗体‎bifun‎c tion‎a l catal‎y st 双官能催化‎剂,双功能催化‎剂bifun‎c tion‎a l initi‎a tor 双官能引发‎剂,双功能引发‎剂bifun‎c tion‎a l inter‎c alat‎o r 双功能嵌人‎剂,双功能插入‎剂bifun‎c tion‎a l linke‎r双功能接头‎bifun‎c tion‎a l vecto‎r双功能载体‎bifur‎c ated‎hydro‎g en bond 分叉氢键bigly‎c an 双糖链蛋白‎聚糖[有两条糖胺‎聚糖链] bibte‎r al symme‎t ry 两侧对称,左右对称bilay‎e r 双层[时特指脂双‎层]bile acid 胆汁酸bile pigme‎n t 胆汁色素bile salt 胆汁盐bile salt micel‎l e 胆汁盐微团‎,胆汁盐胶团‎,胆汁盐胶束‎Bili-[头]胆汁bilin‎胆汁三烯bilin‎o gen 胆汁烷bilin‎o gens‎后胆色素原‎类bilin‎s后胆色素类‎bilir‎u bin 胆红素bilir‎u bin glucu‎r onid‎e胆红素葡糖‎苷酸biliv‎e rdin‎胆绿素bimet‎a llic‎enzym‎e双金属酶bimol‎e cula‎r lipid‎membr‎a ne 双分子脂膜‎bimol‎e cula‎r react‎i on 双分子反应‎bimol‎e cmla‎r reduc‎t ion 双分子还原‎(反应)Binar‎y fissi‎o n 二分裂bindi‎n结合蛋白bindi‎n g site 结合位点,结合部位binoc‎u lar micro‎s cope‎双目显微镜‎binod‎a l curve‎双结点曲线‎binom‎i al distr‎i buti‎o n 二项(式)分布binom‎i al nomen‎c latu‎r e 双名法bioac‎t ive molec‎u le 生物活性分‎子bioac‎t ive pepti‎d e 生物活性肽‎bioac‎t ive polym‎e r 生物活性高‎分子,生物活性聚‎合物bioac‎t ivit‎y生物(学)活性bioam‎i ne 生物胺bioas‎s ay 生物测定,生物学鉴定‎(法)bioas‎t rona‎u tics‎生物航天学‎bioau‎t ogra‎p hy 生物自显影‎(法)BioBe‎a ds resin‎[商] BioBe‎a ds树脂‎[Bio-rad公司‎商品,可用于层析‎及多肽合成‎]BioBr‎e ne solut‎i on [商] BioBr‎e ne溶液‎[由Appl‎i ed Bio-syste‎m s公司生‎产的一种P‎o ly-brene‎类聚卤化季‎铵盐的溶液‎bioca‎b ina 生物舱bioca‎t alys‎i s 生物催化bioca‎t alys‎t生物催化剂‎bioce‎r amic‎生物陶瓷bioch‎e mica‎l生化试剂,生化药品bioch‎e mica‎l engin‎e erin‎g生化工程bioch‎e mica‎l fuel cell 生化燃料电‎池bioch‎e mica‎l mediu‎m生化培养基‎bioch‎e mica‎l mutan‎t生化突变体‎,生化突变型‎bioch‎e mica‎l oxyge‎n deman‎d(BOD) 生化需氧量‎bioch‎e mica‎l polym‎o rphi‎s m 生化多态性‎bioch‎e mist‎r y 生物化学bioch‎i p 生物芯片bioch‎r onom‎e try 生物钟学bioci‎d e 抗微生物剂‎,抗生剂biocl‎i mate‎生物气候biolo‎c k 生物钟bicoe‎n osis‎生物群落bioco‎m muni‎t y(=bioco‎e nosi‎s) 生扬群落bioco‎m pute‎r生物计算机‎bioco‎n trol‎=biolo‎g ical‎contr‎o l)生铂防治;生物控制bioco‎n trol‎syste‎m生物控制系‎统bioco‎n vers‎i on 生物转化[如利用微生物酶‎改造有机化‎合物的分子‎结构]bioco‎s mona‎u tics‎生物宇航学‎biocy‎b erne‎t ics 生物控制论‎biocy‎t in 生物胞素,ε-N-生物素-L-赖氨酸biode‎g rada‎b le (可被)生物降解的‎biode‎g rada‎b le plast‎i cs (可被)生物降解(的)塑料biode‎g rada‎t ion 生物降解(作用)biode‎g rada‎t ion pathw‎a y 生物降解途‎径biode‎t erio‎r atio‎n生物致劣(作用)[生物体的生‎命活动而造‎成物质的物‎理或化学性‎质改变及质量‎降低]biodi‎v ersi‎t y 生物多样性‎Bio-Dotmi‎c rofi‎l trat‎i on appar‎a tus [商] Bio-Dot微置‎过滤装置[Bio-Rad公司‎商品,可用于蛋白‎质或核酸溶‎液的抽滤,并能使样品‎在滤膜上形‎成规则的斑‎点或狭线]bioel‎e ctre‎m istr‎y生物电化学‎bioel‎e ctro‎n ics 生物电子学‎bioen‎e rget‎i cs 生物能学bioen‎g inee‎r ing 生物工程(学)bioet‎h ics 生物伦理学‎[讨论生物学‎研究(如基因工程‎,器官移植等‎)所涉及的伦‎理问题]biofe‎e dbac‎k生物反馈biofi‎l ter 生物滤器Bonde‎d phase‎键合相Bonde‎d-phase‎chrom‎a togr‎a phy键‎合相层析boded‎silic‎a键合硅bonde‎d stati‎o nary‎phase‎键合固定相‎Bondi‎n g orbit‎a l 成键轨道bondi‎n g regio‎n键(合)区bond lengt‎h键长bond momen‎t键矩bond order‎键级bond sreng‎t h 键强度bond valen‎c e 键价bond valen‎c e bond lengt‎h corre‎-latio‎n键长关联‎bone 骨,骨骼bone marro‎w骨髓bone marro‎w ceIl 骨髓细胞bone marro‎w stem cell 骨髓干细胞‎bone marro‎w trans‎p lant‎a ion骨‎髓移植bone morph‎o gene‎t ic prote‎i n(BMP) 骨形态发生‎蛋白,骨形成蛋白‎boost‎e r imuni‎z atio‎n加强免疫borax‎硼砂borde‎t ella‎包特菌属borde‎t ella‎perts‎s is 百日咳杆菌‎boron‎(B) 硼borre‎l idin‎疏螺体素botan‎y植物学bottl‎e neck 瓶颈bott1‎e neck effec‎t瓶颈效应botto‎m ferme‎n tati‎o n 下面发酵botto‎m phase‎下相botto‎m yeast‎下面酵母botu1‎i nus toxin‎肉毒杆菌毒‎素bound‎a ry 边界;界面bound‎a ry eleme‎n t 边界元件bound‎a ry layer‎边界层bound‎a ry lipid‎界面脂′bound‎a ry zone 界面区bound‎auxin‎束缚生长素‎bound‎water‎结合水,束缚水bouqu‎e t stage‎花束期bovin‎e牛bovin‎e leuke‎m ia virus‎(BLV)牛白血病病‎毒bovin‎e pance‎a tic ribon‎u clea‎s e 牛胰RNA‎酶bovin‎e pancr‎e atic‎tryps‎i n inhib‎i ttor‎(BPTI) 牛胰胰蛋白‎酶抑制剂bovin‎e papil‎l omav‎i rus[BPV)牛乳头瘤病‎毒bovin‎e serum‎album‎i n(BSA)牛血清白蛋‎白bovin‎e splee‎n phosp‎h odie‎s tera‎s e 牛脾磷酸二‎酯酶bovin‎e viral‎diarr‎h ea virus‎(BVDV) 牛病毒性腹‎泻病毒[归于黄病毒‎科瘟病毒属‎] Bowma‎n-Birk prote‎a se inhib‎i torBowma‎n—Birk蛋‎白酶抑制剂‎[见于豆科植‎物种子]box 箱;匣,盒;框brach‎i onec‎t in(= tenas‎c in) 臂粘连蛋白‎brady‎[词头] 缓慢brady‎k inin‎缓激肽brady‎k inin‎poten‎t iati‎n g pepti‎d e缓激肽‎增强肽Bragg‎angle‎布拉格角bulld‎o g clamp‎动脉夹bumbl‎e bee venom‎熊蜂毒bumpi‎n g 暴沸,进沸bundl‎e束bundl‎e sheat‎h (维管)束鞘bunga‎r otox‎i n 银环蛇毒紊‎Bunse‎n burne‎r本生灯Bunse‎n cone 本生焰锥Bunse‎n eudio‎m eter‎本生量气管‎Bunse‎n f1ame‎本生焰bunya‎v irus‎布尼乎病毒‎[科名用Bu‎n yavi‎r idae‎]Bunya‎m wera‎virus‎布尼奥罗病‎毒buoya‎n t densi‎t y 浮力密度buoya‎n t densi‎t y centr‎i fuga‎t ion浮‎力密度离心‎buret‎滴定管burie‎d resid‎u es 隐蔽残基Burki‎t t lymph‎o ma 伯基特淋巴‎瘤bursa‎of Fabri‎c ius 法氏囊,腔上囊bursi‎n法氏囊肽burso‎p oiet‎i n(=bursi‎n) 法氏囊生成‎素Ccalci‎t onin‎(CT) 降钙素alciu‎t ouin‎-gene- relat‎e d pepti‎d e(CGRP)降钙素基因‎相关calic‎i um(Ca) 钙calci‎u m bindi‎n g prote‎i n 钙结合蛋白‎(质)calci‎u m bindi‎n g site 钙结合部位‎calci‎u m/calmo‎d ulin‎—depen‎d entprote‎i n kinas‎e依赖(于)钙-钙调蛋白的‎蛋白激酶calci‎u m chann‎e l 钙通道alciu‎m chloH‎d e 氯化钙calci‎u m—depen‎d ent neutr‎a l prote‎a se (=calpa‎i n) 依赖(于)钙的中性蛋‎白酶calci‎u m-depen‎d ent prote‎i n 依(赖于)钙(的)蛋白(质)calci‎u m influ‎x钙流入calci‎u m media‎t ory prote‎i n 钙中介蛋白‎(质)calci‎u m phoqh‎a te 磷酸钙calci‎u m phosp‎h ate prcip‎i tati‎o n磷酸钙‎沉淀calci‎u m phosp‎h ate—DNA copre‎-cipit‎a te 磷酸钙一D‎N A共沉淀‎物calci‎u m phosp‎h ate-DNA copre‎-cipit‎a tion‎磷酸钙→DNA共沉‎淀calci‎u m pump 钙氟calci‎u m senso‎r prote‎i n 钙传感蛋白‎(质)calci‎u m seque‎s trat‎h n 集钙(作用)calcy‎c lin 钙(细胞)周期蛋白calcy‎p hosi‎n e 钙磷蛋白[是依赖于c‎A MP的蛋‎白激酶的磷‎酸化底物]calde‎s mon 钙调(蛋白)结合蛋白[主要见于平‎滑肌,可与钙调蛋‎白及肌动蛋‎白结合]calel‎e ctri‎n(=annex‎i nⅥ) 钙电蛋白[最初发现于‎鳗鱼电器官‎的一种钙结‎合蛋白Ca‎l f intes‎t inal‎alkal‎i ne phosp‎h atas‎e(CIP)(小)牛小肠碱性‎磷酸酶calf serum‎小牛血清calft‎h ymus‎小牛胸腺calgr‎a nuli‎n钙粒蛋白calib‎r atio‎n校准,标准calib‎r atio‎n curve‎校正曲线calib‎r atio‎n filte‎r校准滤光片‎calib‎r atio‎n prote‎i n 校准蛋白calic‎h eamy‎c im 刺孢霉素[来自刺孢小‎单胞菌的抗‎肿瘤抗生素‎,带有二炔烯‎官能团]calic‎i viru‎s杯状病毒[科名用Ca‎l iciv‎i rida‎e]callo‎s e 胼胝质,愈伤葡聚糖‎,capso‎m er(e) (病毒)壳粒capsu‎l ar polys‎a ccha‎r ide 荚膜多糖capsu‎l atio‎n包囊化(作用),胶囊化(作用)capsu‎l e 荚膜capsu‎l e swell‎i ng react‎i on 荚膜肿胀反‎应captu‎r e 捕捉,俘获captu‎r e antig‎e n(=coati‎n g antig‎e n) 捕捉抗原[酶免疫测定‎中用于捕捉‎抗体的抗原‎] captu‎r e assay‎捕捉试验carba‎m yl 氨甲酰基carba‎m ylat‎i on 氨甲酰化carba‎m yl ornit‎h ine 氨甲酰鸟氨‎酸carba‎m ylph‎o spha‎t e 氨甲酰磷酸‎carba‎m yl phosp‎h ate synth‎e tase‎氨甲酰磷酸‎合成酶carba‎m yl trans‎f eras‎e氨甲酰(基)转移酶carba‎n ion 碳负离子carba‎n yl group‎羰基cirbe‎n e 卡宾carbe‎n icil‎l in 羧苄青霉素‎carbe‎n oid 卡宾体carbo‎c atio‎n碳正离子carbo‎d iimi‎d e 碳二亚胺carbo‎h ydra‎t e 糖类,碳水化合物‎carbo‎h ydra‎t e bindi‎n g prote‎i n糖结合‎蛋白carbo‎h ydra‎t e finge‎r prin‎t ing 糖指纹分析‎carbo‎h ydra‎t e mappi‎n g 糖作图,糖定位Carbo‎h ydra‎t e-prote‎i n inter‎a ctti‎o n 糖-蛋白质相互‎作用carbo‎h ydra‎t e seque‎n cing‎糖测序carbo‎l fuchs‎i n 石炭酸品红‎carbo‎l ine 咔啉,二氮芴cirbo‎n(C) 碳carbo‎n assim‎i htio‎n碳同化carbo‎n ate 碳酸盐,碳酸酯carbo‎n ated‎plant‎碳化植物carbo‎n balan‎c e 碳平衡carbo‎n cycli‎n g 碳循环carbo‎n dioxi‎d e 二氧化碳carbo‎n dioxi‎d e compe‎n sati‎o npoi‎n t 二氧化碳补‎偿点carbo‎n dioxi‎d e ferti‎l iati‎o n 二氧化碳施‎肥carbo‎n dioxi‎d e fixat‎i on 二氧化碳固‎定carbo‎n dioxi‎d e tensi‎o n 二氧化碳张‎力carbo‎n fiber‎碳纤维carbo‎n fixat‎h n 碳固定carbo‎n ic anhyd‎r ase 碳酸酐酶carbo‎n isoto‎p e 碳同位素carbo‎n isoto‎p e analy‎s is 碳同位素分‎析carbo‎n isoto‎p e compo‎s itio‎n碳同位素组‎成carbo‎n ium ion 碳正离子carbo‎n monox‎i de 一氧化碳carbo‎n sourc‎e碳源carbo‎n yl 羰基carbo‎n ylat‎i on 羰基化carbo‎x ydis‎m utas‎e羧基歧化酶‎,核酮糖二磷‎酸羧化酶Carbo‎x ydot‎r ophi‎c bacte‎r ia 二氧化碳营‎养菌基质相‎互作用Cell-media‎t ed immun‎i nity‎(CMI)细胞(介导)免疫Cell media‎t ed lymph‎o cyto‎t oxit‎y细胞介导‎的淋巴细胞‎毒性cll membr‎a ne (细)胞膜cll migra‎t ion 细胞迁移cll mobil‎i ty 细胞运动性‎cell motil‎i ty(= cell mobil‎i ty)细胞运动性‎cell movem‎e nt 细胞运动cell mutat‎i on 细胞突变cello‎b iose‎纤维二糖酶‎cello‎i se 纤维二糖cello‎b iuro‎n ic acid 纤维二糖醛‎酸cello‎-oligo‎s acch‎a ride‎纤维寡糖cello‎s olve‎溶纤剂,纤溶剂cello‎t rios‎e纤维三糖cell param‎e ters‎(=latti‎c e param‎e ters‎)晶胞参数cell perme‎a bili‎z ntio‎n细胞透化(作用)[使细胞的通‎透性增加l‎cell polar‎i ty 细胞极性cell recog‎n itio‎n细胞识别cell recyc‎l e 细胞再循环‎[如见于发酵‎工艺]cell repos‎i tory‎(=cell bank) 细胞库cell respi‎r atio‎n细胞呼吸cell ruptu‎r e 细胞破裂,细胞破碎cell rhyth‎m细胞节律cell satte‎r ing facto‎r细胞分散因‎子cell scrap‎e r 细胞刮棒cell senes‎c ence‎(=cell aging‎)细胞衰老cell shape‎细胞形状cell signa‎l ing 细胞信号传‎导,细胞信号发‎放cell socio‎l ogy 细胞社会学‎cell sorte‎r细胞分选仪‎cell sorti‎n g 细胞分选cell strai‎n细胞株[从原代培养‎或细胞系获‎得,具有特定性‎质或标志并‎在晦后培养‎期间始终保‎持]cell subst‎r ain 细胞亚株cell subst‎r ate attac‎h ment‎细胞基底附‎着cell subst‎r ate inter‎a ctio‎n细胞-基底相互作‎用cell surfa‎c e 细胞表面cell surfa‎c e recep‎t or 细胞表面受‎体cell surfa‎c e recog‎n itio‎n细胞表面识‎别cell targe‎t ing 细胞寻靶cell theor‎y细胞学说cell thera‎p y 细胞治疗[如LAK,TIL等方‎法]cell trans‎f omat‎i on 细胞转化cell tropi‎s m 细胞嗜性cell typin‎g细胞分型cellu‎l ar 细胞的;微孔的cellu‎l ar compa‎r tmen‎t细胞区室cellu‎l ar immun‎i ty 细胞免疫cellu‎l ar immun‎o logy‎细胞免疫学‎cellu‎l ar local‎i zati‎o n 细胞定位cellu‎l ar mater‎i al 微孔材料cellu‎l ar oncog‎e ne(c-oncog‎e ne)细胞癌基因‎chemi‎l umin‎o metr‎y化学发光分‎析(法)chemi‎o smos‎i s 化学渗透chemi‎o smot‎i c hypot‎h esis‎化学渗透假‎说chemi‎o smot‎i c theor‎y化学渗透理‎论chemi‎s orpt‎i on 化学吸附chemi‎s try 化学chemo‎- [词头] 化学chemo‎a ttra‎c tant‎化学引诱物‎,化学吸引物‎,趋化物chemo‎a ttra‎c ting‎cytok‎i ne 趋化demil‎u mino‎m etry‎化学发光分‎析(法)chemi‎o smos‎i s 化学渗透chemi‎o smot‎i c hypot‎h esis‎化学渗透假‎说chemi‎o smot‎i c theor‎y化学渗透理‎论chemi‎s orpt‎i on 化学吸附chemi‎s try 化学chemo‎- [词头] 化学chemo‎a ttra‎c tant‎化学引诱物‎,化学吸引物‎,趋化物chemo‎a ttra‎c ting‎cytok‎i ne 趋化chemo‎o rgan‎o trop‎h y 化能有机营‎养chemo‎r ecep‎t ion 化学感受(作用)chemo‎r ecep‎t or 化学感受器‎chemo‎r epel‎l alt 化学排斥物‎chemo‎s enso‎r y neuro‎n感化性神经‎元chemo‎s enso‎r y trans‎d ucer‎感化性传导‎物chemo‎s mosi‎s(=chemi‎o smos‎i s)化学渗透chemo‎s tat 恒化器chemo‎s tat cultu‎r e 恒化培养(物)chemo‎s ynth‎e sis 化能合成chemo‎t acti‎c cytok‎i ne 趋化细胞因‎子chemo‎t acti‎c facto‎r趋化因子chemo‎t acti‎c lipid‎趋化脂质chemo‎t acti‎c pepti‎d e 趋化肽chemo‎t acti‎c repel‎l ant 化学拒斥剂‎chemo‎t axin‎趋化因子,趋化物ch‎e mota‎x is 趋化性[(细胞)随环境中化‎学物质的分‎布梯度作定‎向运动]chemo‎t hera‎p euta‎n t 化疗剂chemo‎t hera‎p euti‎c agent‎化疗剂chemo‎t hera‎p euti‎c s 化疗药物chemo‎t hera‎p y 化疗,化学疗法c‎h emot‎r ophy‎化能营养chemo‎t ropi‎s m 向化性[因环境中化‎学物质的浓‎度梯度而引‎起细胞的定‎向生长]chemo‎t ype 化学型chemo‎v ar 化学变型chrom‎o soma‎l disor‎d er 染色体病chrom‎o soma‎l DNA染色体DN‎Achrom‎o soma‎l domai‎n染色体结构‎域chrom‎o soma‎l elimi‎n atio‎n染色体丢失‎,染色体消减‎chrom‎o soma‎l local‎i zati‎o n 染色体定位‎chrom‎o soma‎I mutat‎i on 染色体突变‎chrom‎o soma‎I patte‎r n 染色体型chrom‎o soma‎l poIym‎o rphi‎s m 染色体多态‎性chrom‎o soma‎l puff 染色体胀泡‎,染色体疏松‎chrom‎o soma‎l rearr‎a ngem‎e nt染色‎体重排chrom‎o soma‎l scaff‎o ld 染色体支架‎chrom‎o some‎染色体chrom‎o some‎aberr‎a tion‎染色体畸变‎chrom‎o some‎abnor‎m alty‎染色体异常‎chrom‎o some‎arm 染色体臂chrom‎o some‎band 染色体带chrom‎o some‎bandi‎n g techn‎i que染‎色体显带技‎术chrom‎o some‎blott‎i ng 染色体印迹‎[通过脉冲电‎场凝胶电泳‎分离染色体‎D NA并进‎行Sout‎h ern印‎迹]chrom‎o some‎break‎a ge 染色体断裂‎chrom‎o some‎bridg‎e染色体桥chrom‎o some‎cente‎r(=chrom‎o cent‎e r) 染色中心chrom‎o some‎coili‎n g 染色体螺旋‎chrom‎o some‎compl‎e ment‎染色体组chrom‎o some‎core 染色体轴chrom‎o some‎crawl‎i ng 染色体缓移‎chrom‎o some‎cycle‎染色体周期‎c hrom‎o some‎damag‎e染色体损伤‎chrom‎o some‎delet‎i on 染色体缺失‎chrom‎o some‎doubb‎l ing 染色体加倍‎chrom‎o some‎dubli‎c atio‎n染色体重复‎chrom‎o some‎elimi‎n atio‎n染色体丢失‎,染色体消减‎chrom‎o some‎engin‎e erin‎g染色体工程‎(学)chrom‎o some‎evolu‎t ion 染色体进化‎chrom‎o some‎excha‎n ge 染色体交换‎chrom‎o some‎flow sorti‎n g 染色体流式‎分选(法)chrom‎o some‎fragm‎e ntat‎i on 染色体断裂‎chrom‎o some‎fusio‎n染色体融合‎chrom‎o some‎gap 染色体间隙‎,染色体裂隙‎chrom‎o some‎inter‎c hang‎e染色体互换‎chrom‎o some‎jumpi‎n g 染色体跳查‎conde‎n ser 冷凝器;聚光器conde‎n sing‎agent‎凝聚剂[大分子群集‎作用的促进‎剂]缩合剂[缩合反应的‎催化剂];冷凝剂condi‎t ion 条件condi‎t iona‎l cytot‎o xic 条件致细胞‎毒condi‎t iona‎l letha‎l条件致死condi‎t iona‎l letha‎l mutan‎t条件致死突‎变体,条件致死突‎变型condi‎t iona‎l letha‎l mutat‎i on 条件致死突‎变condi‎t iona‎l mutan‎t条件突变体‎,条件突变型‎,条件突变株‎condi‎t iona‎l mutat‎i on 条件突变condi‎t ione‎d mediu‎m条件培养基‎,条件培养液‎condi‎t iomd‎patho‎g en 条件致病菌‎,条件致病病‎原体condi‎t ioni‎n g 调节,调理,再生condu‎c tion‎传导condu‎c tivi‎t y 电导率,导电率;导电性;传导率;传导性condu‎c tivi‎t y detec‎t or 电导率检测‎器condu‎c tivi‎t y water‎电导水comdu‎c tome‎t er 电导计condu‎c tome‎t ric titra‎t ion 电导滴定(法)condu‎c tome‎t ry 电导分析(法)cone 锥,圆锥体cone and plate‎visco‎m eter‎锥板式粘度‎计confd‎e nce coeft‎i cien‎t置信系数confd‎e ng inter‎v al 置信区间confi‎d ence‎limit‎置信限confi‎g urat‎i on 构型confg‎u rati‎o nal disor‎d er 构型无序confg‎u rati‎o nal unit 构型单元confi‎r mato‎r y seque‎n cing‎确证性测序‎,证实性测序‎confi‎r mato‎r y test 确证试验,证实试验confl‎u ent 汇合的,融合的,连生的,铺满的confl‎u ent culti‎v atio‎n铺满培养confl‎u ent cultu‎r e 铺满培养物‎,铺满培养confl‎u ent monol‎a yer cells‎(汇合)成片的单层‎细胞,铺满的单层‎细胞confo‎r mati‎o n 构象confo‎r mati‎o nal analy‎s ls 构象分析confo‎r mati‎o nal chang‎e构象变化confo‎r mati‎o nal detem‎i nant‎构象(性)决定簇confo‎r mati‎o nal disor‎d er 构象无序confo‎r mati‎o nal epito‎p e 构象(性)表位confo‎r mati‎o nal inver‎s ion 构象反转confo‎r mati‎o nnl repea‎t ing unit构‎象重复单元‎confo‎r mati‎o nal restr‎i ctio‎n构象限制(性)confo‎r mati‎o nal simil‎a rity‎构象相似性‎confo‎r mati‎o nal switc‎h构象转换(器),构象开关confo‎r mati‎o nal witch‎i ng 构象转换cross‎link 交联;交联键cross‎-linke‎d交联的cross‎-linke‎d gel 交联凝胶cross‎-linke‎d netwo‎r k 交联网络coss-linke‎d polym‎e r(=three‎dimen‎s iona‎l polym‎e r) 交联聚合物‎cross‎l inke‎r交联剂cross‎l inki‎n g 交联crosd‎i nkin‎g agent‎交联剂cross‎l inHn‎g densi‎t y 交联密度cross‎l inki‎n g group‎交联基cross‎l inki‎n g index‎交联指数cross‎match‎i ng 交叉配血(试验)cross‎neutr‎a1iza‎t ion 交叉中和cross‎v er flixa‎t ion交‎换固定cross‎v er suppr‎e ssor‎交换抑制因‎子cross‎v er unit 交换单位cross‎v er value‎交换值coss phosp‎h oryl‎a tion‎交叉磷酸化‎cross‎polli‎n atio‎n异体受粉,异花传粉cross‎prote‎c tion‎交叉保护cross‎-react‎i ng antib‎o dy 交叉反应抗‎体cross‎-react‎i ng antig‎e n 交叉反应抗‎原cross‎-react‎i ng deter‎m inan‎t(CRD) 交叉反应决‎定簇cross‎react‎i on 交叉反应cross‎react‎i vati‎o n 交叉复活cross‎regul‎a tion‎交叉调节,交互调节cross‎secti‎o n 截面,横切面cross‎-steri‎l ity 杂交不育,杂交不育性‎cross‎t alk 串话,通讯crota‎p olin‎(= croto‎x in A) 响尾蛇毒素‎Acroti‎n巴豆毒蛋白‎croto‎x in 响尾蛇毒素‎crown‎gall 冠瘿crown‎gall nodul‎e冠瘿瘤cruci‎b le 坩埚cruci‎f oml 十字形,十字形结构‎cruci‎f oml loop 十字形环[双股反向重‎复序列的每‎一股分别自‎我配对而形‎成] cruci‎f orm struc‎t ure 十字形结构‎crude‎粗的,粗制的crude‎extra‎c t 粗提物crude‎produ‎c t 粗制品crust‎a cyan‎i n 虾青蛋白,甲壳蓝蛋白‎cryo-[词头]冷,冻cryob‎i o1og‎y低温生物学‎cryoc‎o ncen‎t rati‎o n 低温浓缩cryoc‎r ysta‎l1ogr‎a phy 低温晶体学‎cryoe‎l ectr‎o n micro‎s copy‎冷冻电(子显微)镜术cryof‎i xati‎o n 冷冻固定cryog‎e n 冷冻剂,制冷剂cryog‎e nic g1ove‎低温防护手‎套cryog‎e nic purif‎i cati‎o n 深低温纯化‎cryog‎e nic refri‎g erat‎o r 低温冰箱cryog‎e nics‎低温实验法‎cryog‎e nic separ‎a tion‎深低温分离‎cytop‎l asmi‎c regio‎n胞质区[如指跨膜蛋‎白的胞内区‎]cytop‎l asmi‎c strea‎m ing(=cyclo‎s is) 胞质环流cytop‎l asmi‎c tall 胞质尾区[如指跨膜蛋‎白的胞内小‎区]cytop‎l asm IgM 胞质IgM‎,胞质免疫球‎蛋白Mcytop‎l ast 胞质体cytor‎h eolo‎g y 细胞流变学‎cytos‎o l 胞质溶胶cytos‎o lic local‎i zati‎o p. 胞质定位cytos‎o lic space‎胞质腔cytos‎o l recep‎t or 胞质受体cytos‎o me(=cytop‎l ast) 胞质体cytos‎t atic‎facto‎r细胞静止因‎子cytot‎a ctin‎(=tenas‎c in) 腱生蛋白cytot‎o xic 细胞毒的cytot‎o xici‎t y 细胞毒性cytot‎o xic T cell(Tc)(=cytot‎o xic T lymph‎o cyte‎)细胞毒性T‎细胞cytot‎o xic T lymph‎o cyte‎(CTL,Tc) 细胞毒性Υ‎淋巴细胞cytot‎o xin 细胞毒素cytot‎r opis‎m细胞向性cytov‎i llin‎细胞绒毛蛋‎白[见于微绒毛‎,可作用于肌‎动蛋白丝]Ddalto‎n道尔顿[符号D,Da]damma‎r ane 达玛烷damma‎r anet‎y pe 达玛烷型Dane parti‎c le 丹氏粒[型肝炎病毒‎的完整毒粒‎]dange‎r ous 危险的dansy‎l丹(磺)酰,l-二甲氨基萘‎—5-磺酰dansy‎l chlo‎r ide 丹磺酰氯dansy‎l metho‎d丹磺酰法dantr‎o lene‎硝苯呋海因‎[肉松弛剂]dark curre‎n t 暗电流dark field‎暗视野,暗视场dark-field‎micro‎s cope‎暗视野显微镜,暗视场显微‎镜dark-field‎micro‎s copy‎暗视野显微术,暗视场显微‎术dark react‎i on 暗反应dark repai‎r暗修复dark respi‎r atio‎n暗呼吸dark room 暗室,晴房dark seed 需暗种子data(单:datum‎)数据,资料data accum‎u lati‎o n 数据累积data acqui‎s itio‎n数据获取decid‎u ous leaf 落叶decli‎n e phase‎细胞生长曲‎线的衰亡期‎decoa‎g ulan‎t(= antic‎o agul‎a nt)抗凝剂decod‎i ng 译码,解码decom‎p oser‎分解者指具‎有分解动植‎物残体或其‎排泄物能力‎的微生物: decom‎p ress‎i on 降压,减压decon‎d ensa‎t ion 解凝(聚)decon‎t amin‎a nt 净化剂,去污剂decon‎t amin‎a ting‎agent‎(=decon‎t amin‎a nt) 净化剂,去污剂decon‎t amin‎a tion‎净化,去污decor‎i n 核心蛋白聚‎糖[一种基质蛋‎白聚糖,又称为PG‎-40]dedif‎f eren‎t iati‎o n 去分化,脱分化deep∞lony‎深层菌落deep etchi‎n g 深度蚀刻deep refri‎g erat‎i on 深度冷冻deep—shaft‎airli‎f t ferme‎n tor 深井气升式‎发酵罐deep shaft‎syste‎m深井系统dreti‎o lati‎o n 去黄化,脱黄化defec‎t ive inter‎f erin‎g(DI) 缺损干扰defec‎t ive mufan‎t缺损突变体‎,。

Category TermGOTERM_MF_F GO:0030594~neurotransmitter receptor activityGOTERM_CC_F GO:0005576~extracellular regionGOTERM_MF_F GO:0042165~neurotransmitter bindingGOTERM_MF_F GO:0008188~neuropeptide receptor activityGOTERM_MF_F GO:0042923~neuropeptide bindingGOTERM_CC_F GO:0044421~extracellular region partGOTERM_MF_F GO:0032403~protein complex bindingKEGG_PATHWA hsa04080:Neuroactive ligand-receptor interaction GOTERM_CC_F GO:0005615~extracellular spaceGOTERM_CC_F GO:0005886~plasma membraneGOTERM_CC_F GO:0045177~apical part of cellGOTERM_BP_F GO:0042981~regulation of apoptosisGOTERM_BP_F GO:0010033~response to organic substanceGOTERM_BP_F GO:0043067~regulation of programmed cell death GOTERM_BP_F GO:0010941~regulation of cell deathGOTERM_MF_F GO:0005319~lipid transporter activityGOTERM_CC_F GO:0044459~plasma membrane partGOTERM_BP_F GO:0043434~response to peptide hormone stimulusKEGG_PATHWA hsa04960:Aldosterone-regulated sodium reabsorption GOTERM_BP_F GO:0010876~lipid localizationGOTERM_BP_F GO:0006111~regulation of gluconeogenesisGOTERM_BP_F GO:0032868~response to insulin stimulusGOTERM_BP_F GO:0048585~negative regulation of response to stimulus GOTERM_MF_F GO:0005200~structural constituent of cytoskeleton GOTERM_BP_F GO:0060173~limb developmentGOTERM_BP_F GO:0048736~appendage developmentGOTERM_BP_F GO:0051094~positive regulation of developmental process GOTERM_BP_F GO:0001501~skeletal system developmentGOTERM_BP_F GO:0042445~hormone metabolic processGOTERM_BP_F GO:0007155~cell adhesionGOTERM_BP_F GO:0022610~biological adhesionGOTERM_BP_F GO:0009719~response to endogenous stimulusGOTERM_MF_F GO:0005158~insulin receptor bindingGOTERM_BP_F GO:0006869~lipid transportGOTERM_MF_F GO:0008528~peptide receptor activity, G-protein coupled GOTERM_MF_F GO:0001653~peptide receptor activityGOTERM_BP_F GO:0009991~response to extracellular stimulusGOTERM_BP_F GO:0006790~sulfur metabolic processGOTERM_BP_F GO:0040008~regulation of growthGOTERM_BP_F GO:0010817~regulation of hormone levelsGOTERM_BP_F GO:0007610~behaviorGOTERM_BP_F GO:0060429~epithelium developmentGOTERM_BP_F GO:0030326~embryonic limb morphogenesisGOTERM_BP_F GO:0035113~embryonic appendage morphogenesisGOTERM_BP_F GO:0007242~intracellular signaling cascadeGOTERM_BP_F GO:0048598~embryonic morphogenesisGOTERM_BP_F GO:0034754~cellular hormone metabolic processGOTERM_MF_F GO:0043168~anion bindingGOTERM_BP_F GO:0019932~second-messenger-mediated signaling GOTERM_BP_F GO:0031667~response to nutrient levelsGOTERM_BP_F GO:0010906~regulation of glucose metabolic process KEGG_PATHWA hsa00980:Metabolism of xenobiotics by cytochrome P450 GOTERM_BP_F GO:0015698~inorganic anion transportGOTERM_CC_F GO:0005887~integral to plasma membraneGOTERM_MF_F GO:0005496~steroid bindingGOTERM_CC_F GO:0016324~apical plasma membraneGOTERM_BP_F GO:0002366~leukocyte activation during immune responseGOTERM_BP_F GO:0002263~cell activation during immune responseGOTERM_BP_F GO:0019218~regulation of steroid metabolic processGOTERM_BP_F GO:0008202~steroid metabolic processGOTERM_BP_F GO:0045321~leukocyte activationGOTERM_MF_F GO:0015247~aminophospholipid transporter activityGOTERM_MF_F GO:0004012~phospholipid-translocating ATPase activityGOTERM_BP_F GO:0051240~positive regulation of multicellular organismal process GOTERM_MF_F GO:0003700~transcription factor activityGOTERM_BP_F GO:0010675~regulation of cellular carbohydrate metabolic process GOTERM_BP_F GO:0032870~cellular response to hormone stimulusGOTERM_BP_F GO:0035107~appendage morphogenesisGOTERM_BP_F GO:0035108~limb morphogenesisGOTERM_BP_F GO:0006109~regulation of carbohydrate metabolic processGOTERM_BP_F GO:0010627~regulation of protein kinase cascadeGOTERM_BP_F GO:0048568~embryonic organ developmentGOTERM_BP_F GO:0048732~gland developmentGOTERM_CC_F GO:0031226~intrinsic to plasma membraneGOTERM_MF_F GO:0043565~sequence-specific DNA bindingGOTERM_BP_F GO:0006955~immune responseGOTERM_BP_F GO:0050796~regulation of insulin secretionGOTERM_BP_F GO:0030855~epithelial cell differentiationKEGG_PATHWA hsa04115:p53 signaling pathwayGOTERM_BP_F GO:0042493~response to drugGOTERM_BP_F GO:0007584~response to nutrientGOTERM_MF_F GO:0005179~hormone activityGOTERM_MF_F GO:0005275~amine transmembrane transporter activityGOTERM_BP_F GO:0046631~alpha-beta T cell activationGOTERM_BP_F GO:0006952~defense responseGOTERM_MF_F GO:0008509~anion transmembrane transporter activityGOTERM_BP_F GO:0043408~regulation of MAPKKK cascadeGOTERM_BP_F GO:0046626~regulation of insulin receptor signaling pathway GOTERM_BP_F GO:0008206~bile acid metabolic processGOTERM_MF_F GO:0031404~chloride ion bindingGOTERM_BP_F GO:0046890~regulation of lipid biosynthetic processGOTERM_BP_F GO:0046942~carboxylic acid transportGOTERM_BP_F GO:0015849~organic acid transportGOTERM_MF_F GO:0005198~structural molecule activityGOTERM_BP_F GO:0043066~negative regulation of apoptosisGOTERM_BP_F GO:0002791~regulation of peptide secretionGOTERM_BP_F GO:0019216~regulation of lipid metabolic processGOTERM_BP_F GO:0045597~positive regulation of cell differentiationGOTERM_BP_F GO:0043069~negative regulation of programmed cell deathGOTERM_BP_F GO:0030900~forebrain developmentGOTERM_MF_F GO:0050698~proteoglycan sulfotransferase activityGOTERM_BP_F GO:0045927~positive regulation of growthGOTERM_BP_F GO:0060548~negative regulation of cell deathGOTERM_BP_F GO:0043255~regulation of carbohydrate biosynthetic process GOTERM_BP_F GO:0016337~cell-cell adhesionGOTERM_BP_F GO:0015837~amine transportGOTERM_BP_F GO:0035136~forelimb morphogenesisGOTERM_BP_F GO:0009725~response to hormone stimulusGOTERM_BP_F GO:0015718~monocarboxylic acid transportGOTERM_BP_F GO:0032102~negative regulation of response to external stimulus GOTERM_CC_F GO:0034702~ion channel complexGOTERM_BP_F GO:0042127~regulation of cell proliferationGOTERM_MF_F GO:0005184~neuropeptide hormone activityGOTERM_MF_F GO:0008083~growth factor activityGOTERM_BP_F GO:0007398~ectoderm developmentGOTERM_BP_F GO:0050864~regulation of B cell activationGOTERM_BP_F GO:0032101~regulation of response to external stimulus GOTERM_MF_F GO:0042277~peptide bindingGOTERM_BP_F GO:0009954~proximal/distal pattern formationGOTERM_CC_F GO:0034707~chloride channel complexGOTERM_BP_F GO:0001775~cell activationGOTERM_BP_F GO:0006469~negative regulation of protein kinase activity GOTERM_BP_F GO:0048015~phosphoinositide-mediated signalingGOTERM_CC_F GO:0070161~anchoring junctionGOTERM_BP_F GO:0007160~cell-matrix adhesionGOTERM_BP_F GO:0050810~regulation of steroid biosynthetic processGOTERM_BP_F GO:0051048~negative regulation of secretionKEGG_PATHWA hsa04510:Focal adhesionGOTERM_BP_F GO:0033673~negative regulation of kinase activityGOTERM_BP_F GO:0045893~positive regulation of transcription, DNA-dependent GOTERM_BP_F GO:0007586~digestionGOTERM_BP_F GO:0030166~proteoglycan biosynthetic processGOTERM_BP_F GO:0010565~regulation of cellular ketone metabolic process GOTERM_MF_F GO:0030674~protein binding, bridgingGOTERM_BP_F GO:0051254~positive regulation of RNA metabolic process GOTERM_BP_F GO:0008283~cell proliferationGOTERM_CC_F GO:0031224~intrinsic to membraneGOTERM_MF_F GO:0015294~solute:cation symporter activityGOTERM_MF_F GO:0005178~integrin bindingGOTERM_MF_F GO:0015171~amino acid transmembrane transporter activityKEGG_PATHWA hsa05110:Vibrio cholerae infectionGOTERM_BP_F GO:0048562~embryonic organ morphogenesisGOTERM_MF_F GO:0015184~L-cystine transmembrane transporter activity GOTERM_MF_F GO:0004994~somatostatin receptor activityGOTERM_BP_F GO:0051348~negative regulation of transferase activity GOTERM_MF_F GO:0015293~symporter activityGOTERM_MF_F GO:0005548~phospholipid transporter activityGOTERM_BP_F GO:0035295~tube developmentGOTERM_BP_F GO:0040014~regulation of multicellular organism growth GOTERM_BP_F GO:0006821~chloride transportGOTERM_CC_F GO:0005925~focal adhesionGOTERM_CC_F GO:0044437~vacuolar partGOTERM_BP_F GO:0051272~positive regulation of cell motionGOTERM_BP_F GO:0031589~cell-substrate adhesionGOTERM_BP_F GO:0005996~monosaccharide metabolic processGOTERM_CC_F GO:0032839~dendrite cytoplasmGOTERM_BP_F GO:0030879~mammary gland developmentKEGG_PATHWA hsa04910:Insulin signaling pathwayGOTERM_BP_F GO:0009952~anterior/posterior pattern formationGOTERM_MF_F GO:0022803~passive transmembrane transporter activityGOTERM_MF_F GO:0000099~sulfur amino acid transmembrane transporter activityGOTERM_BP_F GO:0008285~negative regulation of cell proliferation GOTERM_BP_F GO:0048537~mucosal-associated lymphoid tissue development GOTERM_BP_F GO:0048541~Peyer's patch developmentGOTERM_BP_F GO:0032148~activation of protein kinase B activity GOTERM_BP_F GO:0045471~response to ethanolCount%PValue Genes List Total P op Hits Pop TotalFold Enrich Bonferroni90.309278 2.18E-04SSTR4, SSTR2239512983 5.5155530.1011154 1.85567 3.28E-04F2RL3, OBP2214201012782 1.6046590.07239590.309278 3.80E-04SSTR4, SSTR223103129835.0871610.16936760.206186 5.65E-04SSTR4, SSTR22340129838.732960.24089360.206186 6.34E-04SSTR4, SSTR22341129838.5199610.26624830 1.0309280.001284MIA, IL19, 21496012782 1.866530.254915110.3780070.002009PDPK1, LAMB223196129833.2674340.625121 130.4467350.002026CCKAR, F2RL9325650852.7765880.208053 230.7903780.002411MIA, LY96, 214685127822.0054980.42467383 2.8522340.002953F2RL3, KCNC2143777127821.3125510.491962100.3436430.003091SLC26A4, MU214179127823.3368140.507811 260.8934710.003409GRIK2, STAT236804135281.8536980.998133 240.8247420.003618STAR, LYN, 236721135281.9080840.998731 260.8934710.003874GRIK2, STAT236812135281.8354350.999208 260.8934710.004061GRIK2, STAT236815135281.8286780.9994460.2061860.004392STAR, ATP1022363129835.5447360.88327153 1.8213060.004614CCKAR, KCNC2142203127821.4369660.653290.3092780.005479LEP, PDPK1,236154135283.3499890.99995950.1718210.006094PDPK1, INS,934150856.6679780.50488790.3092780.00614SOAT2, STAR236157135283.2859760.99998830.1030930.007853LEP, INS, N23681352821.495760.99999970.240550.007916LEP, PDPK1,236100135284.012542170.240550.007916LEP, GRB10,236100135284.012542160.2061860.008658TNNT2, CYLC22374129834.7205190.9856470.240550.009099MSX1, RAX, 236103135283.895672170.240550.009099MSX1, RAX, 236103135283.8956721120.4123710.009414LEP, MYOD1,236278135282.4743321 130.4467350.009834HOXD12, IGF236319135282.336008170.240550.010403LEP, UGT1A6236106135283.7854171220.7560140.010459MIA, ICAM4,23670013528 1.801551 220.7560140.010621MIA, ICAM4,23670113528 1.798981 150.5154640.011321LYN, STAR, 236405135282.123038140.1374570.011792PDPK1, GRB122328129838.3171040.99693880.2749140.013349SOAT2, STAR236145135283.162595170.240550.013507SSTR4, SSTR223114129833.5748960.99868870.240550.013507SSTR4, SSTR223114129833.5748960.998688100.3436430.014702LEP, MUC1, 236220135282.605547170.240550.015105GAL3ST3, GG236115135283.4891671130.4467350.016195MYOD1, SOCS236341135282.185297180.2749140.01636LEP, UGT1A6236151135283.0369291160.5498280.017162CCKAR, PRLH236469135281.9555491 100.3436430.017644PSPN, KRT5,23622713528 2.5252160.2061860.017706MSX1, HOXD123687135283.953244160.2061860.017706MSX1, HOXD123687135283.953244133 1.1340210.017786CCKAR, F2RL2361256135281.5060731120.4123710.018564MSX1, ALDH123630713528 2.240601150.1718210.019177UGT1A6, AKR2365913528 4.857799160.2061860.020689SLC26A4, FX2239212983 3.7969390.999963100.3436430.021541SSTR4, SSTR23623513528 2.439235190.3092780.021594LEP, MUC1, 23619713528 2.618773140.1374570.022508LEP, INS, I2363513528 6.55109150.1718210.022643UGT1A6, AKR93605085 4.5564520.92819660.2061860.022927SLC26A4, FX2369313528 3.698196130 1.0309280.023479KCNC1, CCKA214118812782 1.5083070.995664 50.1718210.023795SOAT2, STAR2236412983 4.5484160.999992 70.240550.02388SLC26A4, MU21413312782 3.143630.996053 40.1374570.024247LYN, TICAM123636135286.3691151 40.1374570.024247LYN, TICAM123636135286.3691151 40.1374570.024247LEP, STAR, 23636135286.3691151 90.3092780.024696LEP, SOAT2,236202135282.5539521 100.3436430.02566DOCK2, SPAC236242135282.3686791 30.1030930.0264ATP10A, ATP223151298311.643950.999998 30.1030930.0264ATP10A, ATP223151298311.643950.999998 100.3436430.026524IL27RA, GRI236244135282.3492641 260.8934710.027175MYOD1, ELF5223975129831.5525260.999999 40.1374570.027937LEP, INS, I23638135286.0338981 70.240550.028558PDPK1, GRB1236133135283.0169491 60.2061860.029056MSX1, HOXD123699135283.4740631 60.2061860.029056MSX1, HOXD123699135283.4740631 40.1374570.029887LEP, INS, I23639135285.8791831 100.3436430.030289LEP, LYN, G23624913528 2.302091 80.2749140.030383ALDH1A3, HO236172135282.6661411 70.240550.03042SLC6A3, ALD236135135282.9722541 30 1.0309280.030497KCNC1, CCKA2141215127821.4747890.999169 180.6185570.031472MYOD1, RAX,22360712983 1.726451 200.6872850.031889GBP6, IL27R236690135281.6615081 40.1374570.031908LEP, SSTR5,23640135285.7322031 70.240550.032361KRT5, UPK1B236137135282.9288631 50.1718210.033932SERPINB5, C936850854.0203980.981124 90.3092780.034702UGT1A6, STA236216135282.3884181 70.240550.03542MUC1, SOAT2236140135282.8661021 60.2061860.037789LEP, PENK, 223108129833.2344291 50.1718210.037825SLC6A3, SLC22374129833.9337661 30.1030930.038294DOCK2, INS,23618135289.5536721 180.6185570.039646SPACA3, IL2236615135281.6777181 70.240550.040545SLC26A4, FX223147129832.7723681 60.2061860.041391LEP, GRIK2,236109135283.1553411 30.1030930.042319GRB10, SOCS23619135289.0508471 30.1030930.042319LEP, STAR, 23619135289.0508471 50.1718210.042789SLC26A4, FX22377129833.7805021 40.1374570.043048LEP, STAR, 23645135285.0952921 70.240550.043265AKR1C4, SLC236147135282.7296211 70.240550.044468AKR1C4, SLC236148135282.7111771 180.6185570.044518GFAP, CLDN6223634129831.6529261 120.4123710.045237CDH13, MSX123635413528 1.943121 40.1374570.045481LEP, SSTR5,23646135284.9845251 60.2061860.045623LEP, STAR, 236112135283.0708231 90.3092780.046272LEP, STAR, 236229135282.2528311 120.4123710.048926CDH13, MSX1236359135281.9160571 70.240550.049487SALL3, CCKA23615213528 2.6398311 20.0687290.05043GAL3ST3, CH22331298338.813151 50.1718210.050468MYOD1, INS,2368013528 3.5826271 120.4123710.050662CDH13, MSX123636013528 1.9107341 30.1030930.050814LEP, INS, N23621135288.1888621 100.3436430.05144CDH8, CDH1323627613528 2.0768851 60.2061860.054838LYN, SLC6A323611813528 2.914681 30.1030930.055271HOXA9, ALX423622135287.8166411120.4123710.055727LEP, UGT1A623636713528 1.87429140.1374570.055872AKR1C4, SLC2365013528 4.585763140.1374570.055872LEP, INS, S2365013528 4.5857631 80.2749140.05613KCNC1, FXYD214205127822.3308870.999998 210.7216490.056536TP53I11, LY236787135281.5295591 30.1030930.058206PENK, IGF1,22323129837.5938781 70.240550.058289LEP, MIA, P223161129832.5312931 80.2749140.058352LAMB3, KRT1236199135282.3044031 40.1374570.058632IL27RA, STA23651135284.4958461 70.240550.059079LEP, MYOD1,236159135282.5236121 80.2749140.059787SSTR4, SSTR223203129832.2943741 30.1030930.059859HOXA10, HOX23623135287.4767871 40.1374570.063673FXYD3, GABR21455127824.3439251 100.3436430.064492DOCK2, SPAC236287135281.9972831 50.1718210.0648CBLC, PDPK12368713528 3.294371 50.1718210.067004F2RL3, CCKA23688135283.2569341 70.240550.067994TNS4, ARHGA21417212782 2.430831 50.1718210.069246MIA, PDPK1,23689135283.2203391 30.1030930.069408LEP, STAR, 23625135286.8786441 40.1374570.070294INS, TRH, K23655135284.1688751 80.2749140.070485PDPK1, LAMB9320150852.1762160.999776 50.1718210.071527CBLC, PDPK123690135283.1845571 140.48110.073482MYOD1, UTF1236477135281.6824081 50.1718210.073846SOAT2, CCKA23691135283.1495621 30.1030930.074357GAL3ST3, IG23626135286.6140811 40.1374570.076487LEP, INS, F23657135284.0225991 50.1718210.07745ARHGAP4, SH22394129833.0967941 140.48110.077868MYOD1, UTF1236481135281.6684171 130.4467350.078951MIA, SOX11,236436135281.7091431 103 3.5395190.079701KCNC1, F2RL214548512782 1.121621 50.1718210.079825SLC6A3, SLC22395129833.0641961 40.1374570.079917ICAM4, LYN,2235912983 3.94711 40.1374570.079917SLC6A6, SLC2235912983 3.94711 40.1374570.080017KDELR2, CFT93565085 3.905530.999932 60.2061860.082235ALDH1A3, HO236133135282.5859561 20.0687290.082634SLC3A1, SLC22351298323.287891 20.0687290.082634SSTR4, SSTR22351298323.287891 50.1718210.086003CBLC, PDPK123696135282.9855231 60.2061860.086016SLC6A3, SLC223137129832.5497691 30.1030930.087449ATP10A, ATP22329129836.0227311 80.2749140.088577TNS3, PSPN,236220135282.0844381 40.1374570.08956LEP, SLC6A323661135283.7588221 40.1374570.08956FXYD3, SLC123661135283.7588221 50.1718210.090722TNS4, ARHGA214102127822.9278911 40.1374570.090919CD1B, SLC3A21464127823.7330611 50.1718210.091123CDH13, LYN,23698135282.924594150.1718210.091123MIA, PDPK1,2369813528 2.924594180.2749140.091661LEP, GAL3ST23622213528 2.065659120.0687290.095928GRIK2, ADA21461278219.90966140.1374570.096424SLC6A3, STA2366313528 3.639494160.2061860.096698CBLC, PDPK1931355085 2.4301080.99999260.2061860.097105HOXD4, GATA23614013528 2.4566591 120.4123710.097719KCNC1, FXYD22341312983 1.691614120.0687290.098327SLC3A1, SLC22361298319.406581110.3780070.098867SSTR4, SSTR23636113528 1.7466551 20.0687290.099823STAT5A, ADA23661352819.107341 20.0687290.099823STAT5A, ADA23661352819.107341 20.0687290.099823PDPK1, INS23661352819.107341 40.1374570.099934UGT1A6, STA23664135283.5826271BenjaminiFDR0.101110.312985 0.0723950.419542 0.0886090.544235 0.0877770.807252 0.0744770.906301 0.136817 1.632758 0.178177 2.844165 0.208053 2.283971 0.168291 3.045419 0.155744 3.717749 0.132187 3.888317 0.998133 5.648121 0.964382 5.985058 0.907493 6.394447 0.846179 6.694047 0.300916 6.121242 0.161802 5.752565 0.867578.929442 0.296358 6.726075 0.8487329.9548110.874112.56065 0.8392712.65639 0.8392712.65639 0.45457311.73161 0.84568914.41237 0.84568914.41237 0.82455614.87464 0.80853615.48624 0.79881116.30995 0.77421316.39046 0.75422516.6227 0.75257617.62194 0.51498315.65308 0.7867820.45138 0.52162717.72979 0.52162717.72979 0.79873222.2892 0.78897922.82763 0.79427124.26952 0.78075724.48558 0.78057325.52647 0.77436526.1461 0.76048926.22545 0.76048926.22545 0.74738926.32854 0.7482127.31526 0.74597828.08446 0.63948725.92044 0.77327230.97879 0.76178531.04273 0.76412132.13175 0.58436222.96278 0.7589132.62540.5403326.24218 0.65643729.22134 0.49935326.62952 0.7670534.15834 0.7670534.15834 0.7670534.15834 0.76256834.67236 0.76529435.76273 0.66311631.88416 0.66311631.88416 0.76654636.72509 0.644532.6585 0.77453638.27121 0.7725638.93899 0.76923339.46982 0.76923339.46982 0.76990140.34639 0.76569140.76557 0.75811840.86349 0.75002940.90207 0.54526932.75244 0.67197436.80361 0.75824242.40802 0.7503342.42676 0.74732842.88399 0.6293432.51192 0.76405245.19173 0.76366745.88172 0.71441942.46801 0.69150942.49939 0.7831748.56256 0.78790249.77953 0.69520844.78815 0.79553251.31058 0.79633252.10741 0.79633252.10741 0.69443746.61256 0.79557252.72446 0.79091752.90634 0.79385353.90424 0.6895247.98001 0.79347954.53175 0.78928154.7293 0.784454.84424 0.78332755.36418 0.79635957.43234 0.79460557.85764 0.7170852.41304 0.79568758.59205 0.79158558.73565 0.7872658.84802 0.7861359.30771 0.8023961.71953 0.80000962.016980.79781562.32829 0.79380562.4267 0.79380562.4267 0.73362652.2922 0.7929462.87396 0.75181157.71037 0.73609757.76379 0.79876864.07198 0.79570964.2534 0.79364664.54166 0.72964458.7174 0.79349565.03866 0.74579956.9527 0.81368767.8587 0.81096368.03841 0.81758869.29695 0.73914259.42996 0.82401670.52953 0.82074670.6166 0.82082371.08925 0.81383456.50736 0.82245671.7353 0.82716372.73165 0.82494672.9135 0.82338673.16673 0.82863774.19883 0.80585368.55407 0.83064274.84767 0.83141175.34599 0.76847565.49526 0.80287269.69636 0.7905669.73976 0.7905669.73976 0.79780161.32573 0.84054976.80077 0.78962770.99732 0.78962770.99732 0.8507278.37023 0.79144872.49446 0.78559973.10672 0.85619379.38419 0.85626779.75944 0.85626779.75944 0.78894670.43071 0.76665470.51248 0.85820480.34287 0.85820480.34287 0.85685280.54014 0.76386972.52784 0.86838882.20633 0.81189668.60206 0.86747982.43333 0.8122677.13887 0.80394177.358730.86967983.00799 0.86956283.31227 0.86956283.31227 0.86956283.31227 0.8670883.34731。

Synthesis and evaluation of novel aromatic substrates and competitive inhibitors of GABA aminotransferaseMichael D.Clift and Richard B.Silverman *Department of Chemistry,Department of Biochemistry,Molecular Biology,and Cell Biology,and the Center for DrugDiscovery and Chemical Biology,Northwestern University,Evanston,IL 60208-3113,USAReceived 18September 2007;revised 15October 2007;accepted 16October 2007Available online 22October 2007Abstract—The design,synthesis,and evaluation of novel c -aminobutyric acid aminotransferase (GABA-AT)inhibitors and inacti-vators can lead to the discovery of new GABA-related therapeutics.To this end,a series of aromatic amino acid compounds was synthesized to aid in the design of new inhibitors and inactivators of GABA-AT.All compounds were tested as competitive inhib-itors of GABA-AT.The amino acids with benzylic amines were also tested as substrates for GABA-AT.It was found that these compounds were all poor competitive inhibitors of GABA-AT,but some were substrates of the enzyme,suggesting their utility as scaffolds for potential GABA-AT mechanism-based puter modeling was used to rationalize the substrate activ-ity of the various compounds.Ó2007Elsevier Ltd.All rights reserved.c -Aminobutyric acid aminotransferase (GABA-AT)cat-alyzes the degradation of c -aminobutyric acid (GABA)to succinic semialdehyde (SSA).Control of GABA levels in the body has numerous therapeutic benefits.Depleted levels of GABA,a major inhibitory neuro-transmitter,1have been shown to cause convulsions.2Raising GABA levels in the brain has an anticonvulsant effect;3–5however,direct GABA administration is not effective,as GABA cannot cross the blood brain bar-rier.6Disruption of GABA levels has also been impli-cated in numerous neurological disorders,such as Alzheimer’s disease,7Parkinson’s disease,8Huntington’s disease,9and senile dementia.10Numerous strategies exist to elevate GABA levels in the brain.The strategy that we have taken involves inhibition or inactivation of GABA-AT,the enzyme that is responsible for the degradation of GABA into an inactive form.To enhance the lipophilicity of GABA analogues for more favorable bioavailability,a series of aromatic com-pounds,inspired by the anticonvulsant drug vigabatrin(1)and by 2,a potent conformationally restricted analogue of 111(Fig.1),was designed,synthesized,and evaluated.The results of these studies will aid in the design of future GABA-AT inhibitors and pound 2is a potent irreversible inactivator of GABA-AT,showing a potency that is 186times greater than that of 1.The syntheses of the three 1,3-disubstituted aromatic amino acids (8,10,and 14),that were not commercially available,are outlined in Scheme 1.0960-894X/$-see front matter Ó2007Elsevier Ltd.All rights reserved.doi:10.1016/j.bmcl.2007.10.060Keywords :c -Aminobutyric acid;c -Aminobutyric acid aminotransfer-ase;GABA-AT;Substrate of GABA-AT;Inhibitor;Computer mod-eling;Aromatic amino acid.*Corresponding author.Tel.:+18474915653;fax:+18474917713;e-mail:Agman@Available online at Bioorganic &Medicinal Chemistry Letters 18(2008)3122–3125The commercially unavailable 1,2-disubstituted aro-matic amino acids (16,20,and 25)were synthesized as shown in Scheme 2.The chemistry applied to the synthesis of intermediate 6proved applicable to the synthesis of amino acid 25,but different synthetic strategies were required in the cases of 16and 20.All of the compounds were tested as competitive inhib-itors of GABA-AT using a coupled enzyme assay,12ex-cept for pound 16underwent cyclization to 15under the conditions of the assays and was,therefore,omitted from testing.It was found that 8,10,11,14,20,25,and 26(Fig.2)all showed IC 50values that ex-N H O H 2N .HCl1516NCCO 2HNCCO 2MeN H OCO 2HH 2NCO 2H .HCl17181920CO 2Me .HClCO 2MeBrCO 2MeN 3HNO H 2NCO 2H212223242524abcadefaScheme 2.Synthesis of the 1,2-disubstituted aromatic amino acids.Reagents and conditions:(a)6M HCl,reflux;(b)MeI,NaHCO 3,DMF;(c)H 2,Pd–C,AcOH/MeOH (1:1),then TEA,MeOH;(d)NBS,h m ,MeCN;(e)NaN 3,EtOH,reflux;(f)H 2,Pd–C,MeOH,then TEA,MeOH.M.D.Clift,R.B.Silverman /Bioorg.Med.Chem.Lett.18(2008)3122–31253123ceeded 5.0mM concentrations.Because of the relatively poor potency of these compounds,the exact IC 50values were not determined.All of the compounds were subsequently tested as sub-strates for GABA-AT.The substrate activities (relative to the rate of GABA turnover)for all of the com-pounds are shown in Table 1.The fact that 11,14,and 26showed no transamination was anticipated,as these compounds lack protons alpha to the amino group and,therefore,cannot undergo oxidation at the necessary position.It is clear that the two best substrates are 8and 25.This most likely results from their similarity to the structure of bound GABA in terms of the relative positions of the acid and amine functionalities.The automated flexible docking pro-gram FlexX was employed to perform docking calcu-lations for 8,10,20,and 25using the crystallographic structure of the homodimers of pig liver GABA-AT in complex with vigabatrin 1(PDB code:1OHW).13The FlexX docking models show that none of carboxylic groups of these compounds can exactly mimic the binding mode of the carboxylic group of vigabatrin,as illustrated in Fig.3.However,compared with 10and 20,the carboxylic groups of 8and 25are more similar to that of bound vigabatrin.The poor turn-over of 10may result from the fact that there are too many carbon atoms between the polar groups and,therefore,the carboxylic acid group of 10cannot be accommodated by the active site of GABA-AT (Fig.3a).Table 1.Substrate activities of compounds shown in Fig.2relative to GABA Compound Relative substrate activity (%)GABA 100.008 3.42(±0.82)100.78(±0.06)110.00140.00200.00a25 5.65(±0.86)260.00Numbers represent relative rates of turnover.Standard deviations are also shown.aTransamination was observed,but it was found to be insignificant..HClH 2N CO 2H 25H 2N CO 2H.HCl20.HClNH 2CO 2H10CO 2HNH 2.HCl8H 2NCO 2H11H 2N CO 2H.HCl14H 2NCO 2H26Figure 2.Amino acids that were tested as inhibitors and substrates of GABA-AT.Figure 3.(a)The superimposition of the FlexX docking conformations of 8,10,and 25and the binding conformation of vigabatrin from the crystallographic structure (PDB code:1OHW).The 8–PLP adduct,the 10–PLP adduct,and the 25–PLP adduct are shown in green,red,and purple,respectively.(b)The superimposition of the FlexX docking conformations of 20and the binding conformation of vigabatrin from the crystallographic structure (PDB code:1OHW).Lys329is shown in green.Phe351and Thr353,which are from the other monomer of the homodimer,are shown in magenta.All of the hydrogen atoms were omitted for clarity.Vigabatrin and PLP from the crystal structure are shown in cyan.The 20–PLP adduct is shown in gray.3124M.D.Clift,R.B.Silverman /Bioorg.Med.Chem.Lett.18(2008)3122–3125Compound20has the same chain length as GABA so it is surprising that it lacks substrate proficiency.Docking analysis indicates that the rigidity of the aromatic ring prevents the carboxylate from engaging in a critical salt bridge interaction with Arg-12913present in the active site(Fig.3b).This could explain why25undergoes transamination and20does not;the additional methy-lene unit in25allows the carboxylate to assume an appropriate conformation that facilitates binding by GABA-AT and subsequent turnover.14The results described here suggest that the aromatic scaf-fold of these GABA analogues may be useful in the design of potential mechanism-based inactivators of GABA-AT having increased lipophilicity.It appears that compounds with the general structures related to8and25show the most promise for further investigations.AcknowledgementsThe authors are grateful to the National Institutes of Health(GM66132)forfinancial support of this re-search.We thank Dr.Haitao Ji for carrying out the computer modeling and Dr.Graham Lawton and Dr. Wenxin Gu for helpful discussions.Supplementary data Supplementary data associated with this article can be found,in the online version,at doi:10.1016/j.bmcl. 2007.10.060.References and notes1.Krnjevic,K.Physiol.Rev.1974,54,418.2.Karlsson, A.;Fonnum, F.;Malthe-Sorrensen, D.;Storm-Mathisen,J.Biochem.Pharmacol.1974,23, 3053.3.Gale,K.Epilepsia1989,30,S1.4.Hayashi,T.J.Med.Chem.1981,24,1377.5.Van Gelder,N.M.;Elliott,K.A.C.J.Neurochem.1958,3,139.6.Meldrum,B.S.;Horton,R.W.In Epliepsy;Harris,P.,Mawdsley, C.,Eds.;Churchill Livingston:Edinburgh, 1974.7.Sherif, F.M.;Ahmed,S.S.Clin.Biochem.1995,28,145.8.de Jong,P.J.;Lakke,J.P.;Teelken,A.W.Adv.Neurol.1984,40,427.9.Wu,J.Y.;Bird, E. D.;Chen,M.S.;Huang,W.M.Neurochem.Res.1979,4,475.10.Mountjoy,C.Q.;Rossor,M.N.;Iversen,L.L.;Roth,M.Brain1984,107,507.11.Pan,Y.;Qiu,J.;Silverman,R.B.J.Med.Chem.2003,46,5292.12.Scott,E.M.;Jakoby,W.B.J.Biol.Chem.1959,234,932.13.(a)Storici,P.;Capitani,G.;De Biase,D.;Moser,M.;John,R.A.;Jansonius,J.N.;Schirmer,T.Biochemistry 1999,38,8628–8634;(b)Storici,P.;De Biase,D.;Bossa,F.;Bruno,S.;Mozzarelli,A.;Peneff,C.;Silverman,R.B.;Schirmer,T.J.Biol.Chem.2004,279,363–373.14.The3D coordinates of the active site of GABA-AT(PDBcode:1OHW)in complex with the docking ligands(the8–PLP adduct,the10–PLP adduct,the20–PLP adduct,and the25–PLP adduct)and the crystallographic vigabatrin–PLP adduct are accessible from the supporting information.M.D.Clift,R.B.Silverman/Bioorg.Med.Chem.Lett.18(2008)3122–31253125。

2024 年 3月第 61 卷第 2 期Mar. 2024Vol. 61 No. 2四川大学学报（自然科学版）Journal of Sichuan University （Natural Science Edition）ASI基因家族结构及表达分析黄斌翰，何艳艳，唐婕，叶春，周嘉裕，廖海（西南交通大学生命科学与工程学院，成都 610031）摘要: 为研究编码α-淀粉酶/枯草杆菌蛋白酶抑制剂（α-amylase/subtilisin inhibitor， ASI）的基因结构特征及功能，本研究利用NCBI、Phytozome等平台工具鉴定到来源于23种植物的33个ASI基因家族成员，分析其进化特点及其编码蛋白的理化性质和保守基序等信息.同时结合RT-qPCR技术分析水稻ASI在干旱和盐胁迫条件下的表达模式.结果表明，ASI基因家族具有较高的保守性，但其编码蛋白上下游区域的保守性强弱不一，同时顺式作用元件预测结果提示ASI基因可能参与植物生长发育调控、响应生物及非生物胁迫等生理过程.RT-qPCR分析发现，较未胁迫条件下，幼苗期水稻的根、叶鞘和叶片等组织中RASI基因发生了不同的表达变化，提示ASI基因可能具有多样性的生物学功能.关键词: α-淀粉酶/枯草杆菌蛋白酶抑制剂（ASI）； Kunitz型抑制剂；基因家族鉴定；基因序列分析；基因表达分析中图分类号: Q943.2 文献标志码: A DOI:10.19907/j.0490-6756.2024.026003Structural and expression analysis of the ASI gene familyHUANG Bin-Han， HE Yan-Yan， TANG Jie， YE Chun， ZHOU Jia-Yu， LIAO Hai（School of Life Science and Engineering， Southwest Jiaotong University， Chengdu 610031， China）Abstract: To study the structural features and functions of genes encoding α-amylase/subtilisin inhibitor （ASI），a total of 33 ASI gene family members from 23 plant species were identified in this study by using NCBI，Phytozome and other platforms.The evolutionary characteristics the physicochemical properties and conserved motifs of ASI proteins were analyzed.At the same time， RT-qPCR was combined to analyze the expression pattern of RASI under drought and salinity stress.The results showed that the ASI gene family is highly conserved， but the upstream and downstream regions of the encoded proteins are differently conserved. Meanwhile，the cis-acting element prediction results suggested that the ASI genes may be involved in the physiological processes such as the regulation of plant growth and development，and the response to biotic and abiotic stresses.RT-qPCR analysis revealed that the expression of RASI in roots， leaf sheaths and leaves of rice at the seedling stage changed differently compared with the unstressed condition.These results indicate that ASI genes may have diverse biological functions.Keywords: α-amylase/subtilisin inhibitor， ASI； Kunitz-domain inhibitors； Gene family identification； Gene sequences analysis； Gene expression analysis收稿日期: 2023-04-25基金项目: 国家自然科学基金（32270410）；成都市科技局项目（2022-YF05-01357-SN）；中央高校医工结合项目(2682021ZTPY017，2682022ZTPY053)作者简介: 黄斌翰（1999-），男，四川渠县人，硕士研究生，主要研究领域为分子生物学.E-mail: binhanhuang@ 通讯作者: 廖海.E-mail: ddliaohai@第 2 期黄斌翰，等：ASI基因家族结构及表达分析第 61 卷1　引言α-淀粉酶/枯草杆菌蛋白酶抑制剂（α-amylase/ subtilisin inhibitor，ASI）是具有Kunitz结构域的活性肽，通常含有180~200个氨基酸残基，并含有2个保守的二硫键.ASI的晶体结构呈现β-三叶草形状，其抑制中心突出于三维结构的表面，能够深入到靶酶（α-淀粉酶与枯草杆菌蛋白酶）的活性中心形成靶酶-抑制剂复合物，从而导致靶酶活性的降低［1］.ASI的分子结构中抑制淀粉酶的活性区域分布于C末端，其主要的活性氨基酸残基为谷氨酸［2］，同时精氨酸对于接触靶酶也具有重要作用［3， 4］；而抑制枯草杆菌蛋白酶的活性区域位于肽链中段，主要由苏氨酸发挥抑制作用［5］.根据Raw‑lings等的分类，ASI属于I3抑制剂家族，其主要功能是调控胞外及胞内靶蛋白酶的活性，广泛参与植物生长发育、胁迫应答与病虫害防御等生理过程［6］.例如，ASI不仅能够抑制昆虫体内的α-淀粉酶活性，影响昆虫吸收营养，发挥抗虫的作用；还能够抑制人体肠道内α-淀粉酶的活性，减少糖在人体内的消化吸收，具有降血糖、降血脂的功效.ASI 对微生物来源的枯草杆菌蛋白酶发挥抑制作用，起抗病原菌的作用.此外，干旱、低温等胁迫会明显提高川芎中ASI的mRNA含量，表明其在响应非生物胁迫过程中发挥着重要作用［7］.ASI在被子植物中广泛存在，如在水稻、大麦、小麦与川芎等10余种植物［7-10］中均能找到其存在的证据，但在低等植物如裸子植物与藻类中未发现ASI的存在.为揭示植物ASI的功能与进化特点，有必要利用植物基因组数据开展全基因组鉴定与生物信息学分析.本研究利用在线数据库NCBI、Phytozome及在线分析工具InterPro、SMART鉴定等（种）植物的ASI基因家族成员，使用生物信息学的方法，构建ASI基因家族的系统进化树，并分析ASI基因家族所编码蛋白质的保守序列，此外，通过对实时荧光定量PCR分析水稻ASI的器官特异性及在不同胁迫条件下的表达特征，为进一步分离和克隆植物中的ASI奠定基础.2　材料与方法2.1　材料水稻“中花11号”栽培于中国科学院成都生物研究所.植物总RNA提取试剂TRNzol Universal Reagent购自天根生化科技（北京）有限公司，Swe‑Script All-in-One RT SuperMix反转录试剂盒购自武汉塞维尔生物科技有限公司，TB Green® Pre‑mix Ex Taq™ Ⅱ实时荧光定量试剂盒购自宝日医生物技术（北京）有限公司，其余试剂均为分析纯.2.2　方法2.2.1　ASI基因家族的鉴定及理化性质分析　通过数据库NCBI（https：//www.ncbi.nlm.nih. gov/）和Phytozome（https：//phytozome-next.jgi. /）获得植物来源α-淀粉酶/枯草杆菌蛋白酶抑制剂蛋白序列.初筛去除冗余以及报道的CDS区mRNA与所翻译的蛋白不一致的序列；然后保留其中分子量介于18~22 kD且包含Kunitz 型蛋白特征片段（［LIVM］-x-D-x-［EDNTY］-［DG］-［RKHDENQ］-x-［LIVM］-（x）5-Y-x-［LIVM］）的序列，并获得其genome、GFF、GTF文件.将筛选后的蛋白序列上传至ProtParam （https：///protparam/）线上平台进行理化性质及氨基酸组成分析.同时将通过线上平台SignalP-5.0（https：//services.healthtech. dtu.dk/services/SignalP-5.0/）和PSORT （https：///psort.html），设置默认参数，分别进行信号肽及亚细胞定位预测. 2.2.2　系统发育分析　将鉴定后的ASI基因CDS区蛋白序列使用Clustal W的默认参数进行多重序列比对.然后通过MEGA Ⅹ软件，使用邻近法（Neighbor-Joining Tree），选择Poisson模型，采用Pairwise deletion方法处理Gaps/Missing Date 构建进化树.其可靠性评估采用Bootstrap自展检验法，检验次数为1000次.同时采用极大似然法（Maximum Likelihood Tree）进行验证，选择Jones-Taylor-Thornton（JTT）模型，采用Complete dele‑tion方法处理Gaps/Missing Date构建进化树，其可靠性评估同邻近法.并且参考被子植物系统发育研究组建立的被子植物分类系统第四版（APG Ⅳ，https：//）所确定的物种亲缘关系，编写newick文件展示相应物种进化树. 2.2.3　多序列比对及序列变异分析　通过DNAMAN软件将LASI序列同已有研究报道的其他物种ASI序列进行多重比对分析，采用默认参数，确定保守区域及关键活性位点.将LASI氨基酸序列分别同来源于单子叶植物、双子叶植物第 61 卷四川大学学报（自然科学版）第 2 期及ASI序列进行多重序列比对，比对文件提交至PVS线上工具（http：//imed.med.ucm.es/ PVS/），选择Shannon法［11，12］进行蛋白序列变异分析，输出参数设置默认.2.2.4　保守基序及顺式作用元件分析　利用在线工具MEME（https：///meme/tools/ meme）识别ASI序列中所包含的保守基序，设置最佳Motif数量为8，其余参数设置默认值.同使用TBtools［13］提取所有ASI基因上游2000 bp区域的启动子序列，利用PlantCARE（https：//bioinfor‑matics.psb.ugent.be/webtools/plantcare/html/）进行顺式作用元件分析［14］.2.2.5　非生物胁迫下水稻RASI基因的表达分析　将生长1 w的水稻幼苗置于低浓度的9% PEG和60 mmol/L NaCl胁迫24 h之后，再采用18% PEG 和120 mmol NaCl模拟干旱、盐胁迫培养1 w.收集对照和胁迫后的水稻根、叶鞘及叶片组织，提取总RNA，逆转录后得到cDNA作为模板，利用Primer Premier 5设计引物，交由北京擎科生物有限公司合成（表1），以水稻eEF-1α为内参，通过实时荧光定量PCR分析水稻RASI基因在不同处理条件下在不同组织中的表达情况.3　结果与分析3.1　Kunitz型ASI基因家族的鉴定及理化性质分析通过NCBI等数据库，本研究鉴定到33条植物来源的Kunitz型ASI序列（表2），包括川芎（Li⁃gusticum chuanxiong）、橡胶树（Hevea brasiliensis）、川桑（Morus notabilis）等13种双子叶植物来源的21条，以及水稻（Oryza sativa Japonica Group）、大麦（Hordeum vulgare）等11种单子叶植物来源的12条.33条ASI编码蛋白质由180~234个氨基酸残基组成，平均长度为204 aa，平均分子量约为22.3 kD.理论等电点为4.43~9.6，稳定性差异较大.GRAVY值介于−0.476~0.112之间，大部分呈现负值（81.8%），表现出亲水性（比如水稻O.sativa Japonica Group），此外有6条ASI表现出疏水性且均来源于双子叶植物（如川芎L.chuanx⁃iong为0.083）.通过亚细胞定位预测，发现ASI主要定位在质外体，占54.5%（包括川芎L.chuanx⁃iong、水稻O.sativa Japonica Group等），较少ASI 基因在线粒体或内质网中表达（比如大麦H.vul⁃gare），极少数在高尔基体或细胞质中表达（比如川桑M.notabilis2）.使用SignalP-5.0线上工具对33条ASI进行了信号肽预测，其中30条ASI序列检测到信号肽段，长度范围为18~30 aa，而杨梅M. rubra、川桑M.notabilis2和小麦T.aestivum三条序列未识别到明显信号肽区域.表2　33条ASI序列理化性质、亚细胞定位及信号肽预测Tab.2　Characteristic features subcellular localization and signal peptides of 33 ASI protein sequences黄花蒿黄花蒿黄花蒿木豆木豆柠檬甜橙寡头二蕊草寡头二蕊草大豆野大豆野大豆A.annua1A.annua2A.annua3C.cajan1C.cajan2C.clementinaC.sinensisD.oligosanthes1D.oligosanthes2G.maxG.soja1G.soja2PWA35940.1PWA76553.1PWA88366.1XP_020210107.1XP_020227907.1XP_006448699.1KAH9795663.1OEL21755.1OEL26692.1KAH1210237.1KHM98864.1RZB77313.121620020620119820019521421020620319823.3922.0922.622.6221.821.8521.3523.0523.1623.5422.8321.618.29.29.64.775.535.466.588.46.154.744.434.830.045−0.178−0.217−0.351−0.218−0.009−0.027−0.106−0.114−0.443−0.203−0.073MitochondrialMitochondrialERExtracellularExtracellularERERMitochondrialExtracellularExtracellularGolgiExtracellularMKESFFIFTVLSTLAFSLCMRESIFIFIVLSILTLSSCMKLNILIPTLLSTFLSLYVASSMSTRSIETTLTLMLCLAIIATTALAMSMKLWASLTLAVWLITATSSLAMRQSAMVRLVKSLSFFCLVLAITQAMVRLIKSRSFFCLVLAITQAMDHHHPLLLLLLPLLAISFPCIAMGGRPGAAARLILLLLSAVLAVSLIQRGAAMSMKLYASLTFVVWLFMATSSLAMSMKLLLASLSISVWLFMATLSLAMSMRSIGTSLSLMLWLVIATSALAAnnotation Name Species Name Accession NO.Length（aa）MW（kD）pI GRAVYSubcellularLocalizationSignal Peptides 表1　引物序列信息Tab.1　Information of peimers sequencePrimers RASI-1Sequence （5′→3′）F：TCCGCATCCGCTTCAACGC R：CACGAGTCCCTGCACGACACCA第 2 期黄斌翰，等： ASI 基因家族结构及表达分析第 61 卷野大豆橡胶树大麦大麦（驯化）川芎杨梅川桑川桑水稻稷稷柳枝稷毛果杨毛果杨簸箕柳粱高粱密花豆小麦乌拉尔图小麦玉米G.soja 3H.brasiliensis H.vulgare H.vulgare subsp.vulgare L.chuanxiong M.rubra M.notabilis 1M.notabilis2O.sativa Ja⁃ponica Group iaceum iaceum 2P.virgatum P.trichocarpa 1P.trichocarpa 2S.suchowensis S.italica S.bicolor S.suberectus T.aestivum T.urartu Z.maysRZB98461.1AJG01567.1P07596.2XP_044970355.1AOO34766.1KAB1216374.1EXB74706.1XP_010098224.1P29421.2RLM65953.1RLM75425.1XP_039821064.1XP_002309883.1XP_006371005.1KAG5224050.1XP_004976298.3XP_002448209.1TKY49253.1P16347.1EMS66656.1XP_008669134.120819620320420319019120720021221321319620019621323019718023422223.3721.1622.1622.2522.5720.6820.8222.6521.4222.622.6122.6520.7121.8121.3822.7423.9421.5119.6326.0223.45.265.557.777.775.064.844.468.358.666.546.798.434.584.896.296.358.329.086.777.67.73−0.3150.034−0.469−0.3110.083−0.159−0.09−0.065−0.156−0.118−0.016−0.0530.1120.0530.034−0.026−0.062−0.094−0.476−0.205−0.173Extracellular Extracellular MitochondrialER Extracellular Cytoplasmic Extracellular Cytoplasmic ExtracellularExtracellular Mitochondrial Extracellular Extracellular Extracellular Extracellular Extracellular Extracellular MitochondrialER Mitochondrial ExtracellularMSMKLYASLTLTVWLFMATFSRA MLKLIGSLSFVWLLMAMSTVA MGSRRAGSSSSPLFWPAPPSRA MGSRRAGLLLLSLILASTALSRS MKKNLFYISFLLVALSTYSLVSG ——MNMKYLIGILMSCIWLSMAINA ——MVSLRLPLILLSLLAISFSCSAMGGPGAVPARLLLLLLLSVLAAVSVPR GAAMGGPGAVRLLLLLSVVLAASLYRGAA MGGVPGALRRHILLLLSVALAAISLYRG AAMLRLIGSLSFIWLLMVISTAAQ MLGLIKGLSFICLLMAVSCLA MLRMIGSLSFIWLLMAMSTAAMGGPAGAVRLLLLLLSVLAASLSRVAA MSTKLIGTCLSLMVWLVIATAALA MSSRRVGLLLLSLLATTLTCSA ——MEHFCFLVILSLSGLAMAMPMSIPRAAHLLVLLSVLAISLSSCGAA AA(续表2)Annotation Name Species Name Accession NO.Length （aa ）MW（kD ）pI GRAVY Subcellular Localization Signal Peptides33条ASI 中缬氨酸（Val ）、苏氨酸（Thr ）、脯氨酸（Pro ）、亮氨酸（Leu ）、甘氨酸（Gly ）、精氨酸（Arg ）、丙氨酸（Ala ）及丝氨酸（Ser ）8种氨基酸含量最为丰富，合计比例超过61%（图1）.图1　ASI 主要氨基酸含量Fig.1　Main amino acid contents of ASI proteins第 61 卷四川大学学报（自然科学版）第 2 期3.2　系统发育分析将上述鉴定到的33条ASI 序列以极大似然法（ML ）、邻近法（NJ ）构建系统进化树（图2a 、2b ），在ML 树中超过86%的节点Bootstrap 值大于50，在NJ 树中所有节点Bootstrap 值均大于50，结果可信.同时基于APG Ⅳ系统构建相应的24个物种的系统发育树（图2c ）.两种ASI 进化树与物种树总体上高度相似，表明ASI 进化相对保守.单子叶植物与双子叶植物的ASI 序列分别聚类，形成两个亚家族.川芎LASI （L.chuanxiong ）与黄花蒿ASI （A.annua 1、A.annua 2）亲缘关系最为接近，提示这两类物种ASI 基因生物学功能更为接近.其中A.annua 1与A.annua 2作为黄花蒿并系同源基因聚类在一起，类似情况还有来源于川桑的M.no⁃tabilis 1、M.notabilis 2以及来源于稷的i⁃aceum 1、iaceum 2.此外，ASI 进化树中个别序列聚类位置与物种树存在差异，如在双子叶植物中毛果杨（P.tricho⁃carpa ）与簸箕柳（S.suchowensis ）物种亲缘关系最近，但在两种ASI 进化树中毛果杨P.trichocarpa 2并未与簸箕柳S.suchowensis 聚为一支.类似的情况还有牧豆G.soja 2.而在单子叶植物中，乌拉尔图小麦（T.urartu ）与小麦（T.aestivum ）之间、寡头二蕊草（D.oligosanthes ）与粱（S.italica ）等物种之间亲缘关系更近.但在ASI 进化树中乌拉尔图小麦T.urartu 与寡头二蕊草D.oligosanthes1聚类在一起，并且具有较高的自展值（Bootstrap 值92）；而寡头二蕊草D.oligosanthes 2仍然与粱S.italica 等单子叶植物ASI 聚类在一起.3.3　氨基酸序列比对及蛋白序列变异分析取川芎LASI 、水稻RASI 分别代表双子叶、单子叶植物，与另外5条已有功能研究报道的ASI 序列进行比对分析（图3）.发现不同植物来源的ASI 含有数量不等的Cys 残基，其中靠近N -端的2个Cys 残基（如水稻RASI 的Cys 63、Cys 112残基）较为保守，推测两者之间形成的二硫键，有助于形成对抗枯草杆菌蛋白酶的抑制活性环（水稻RASI 的Ala 108~Ile 111）；而位于下游的Cys 残基保守性较差，来源单子叶植物的ASI 下游含有2处Cys 残基（如RASI 含有Cys 162、Cys 166残基），而双子叶植物的ASI 均含有4处Cys 残基（如川芎LASI 含有Cys 164、Cys 168、Cys 172、Cys 175残基）.图2　ASI 进化树及相应物种进化树Fig.2　ASI phylogenetic tree and taxonomic tree第 2 期黄斌翰，等： ASI 基因家族结构及表达分析第 61 卷通过PVR 线上平台进行蛋白序列变异分析，计算川芎LASI 与其他双子叶植物和单子叶植物ASI 序列的香农熵（图4）.发现ASI 序列N -端信号肽表现出较强变异性，单子叶植物和双子叶植物ASI 序列的平均香农熵分别为1.28和1.51，所有ASI 序列平均香农熵为1.73，说明在双子叶植物中ASI 具有更大变异性.3.4　保守基序分析通过蛋白质保守结构域鉴定，共预测到了8个保守基序（图5），其中基序1在33个ASI 中均有存在，基序2、4仅分别在1条序列中出现缺失（T.uratu 缺失基序2，M.notabilis 1缺失基序4）.绝大部分单子叶植物来源ASI 序列中预测到了基序6（仅D.oligosanthes 1出现缺失），而双子叶植物来源ASI 中均不含有.D.oligosanthes 1与T.urartu 两条ASI 在基序分布上与其他ASI 存在较大差异，二者均无基序3、5、7，T.urartu 还缺失基序8.除此之外，缺失基序8的ASI 序列还包括M.notabilis 1、S.suchowensis 和L.chuanxiong .3.5　顺式作用元件分析ASI 基因成员的顺式作用元件分析如图6所示（因缺失杨梅等8种植物基因组注释文件，故无相应ASI 基因的顺式作用元件分析）.其启动子区域主要含有光（Light ）响应、脱落酸（Abscisic acid ）响应和茉莉酸甲酯（MeJA ）响应元件.同时部分ASI 基因成员还含有生长素（Auxin ）响应、赤霉素（Gibberellin ）响应、水杨酸（Salicylic acid ）响应元件以及参与干旱（Drought ）、低温（Low -temperature ）等防御和应激反应元件.以上结果提示ASI 成员可能具有差异化的生物学功能，并影响植物生长发育、激素应答及逆境响应等过程.3.6　非生物胁迫下水稻RASI 基因的表达分析在无胁迫和干旱、盐胁迫条件处理下，水稻RASI 基因在根、叶鞘和叶片组织中的RT -qPCR 分析结果如图7所示.无胁迫处理的叶鞘组织中，RASI 的表达量较根和叶片出现显著差异，表明图3　7条ASI 氨基酸序列比对图中标记了已知的ASI 结构特征，双向箭头为Kunitz 型蛋白特征序列；圆头箭头为二硫键；倒三角标记枯草杆菌蛋白酶关键结合位点，其方框为活性环［5］；六角星标记α-淀粉酶结合关键位点；LASI （L.chuanxiong ），BASI （H.vulgare ），RASI （O.sativa Japonica Group ），WASI （T.aestivum ），HbASI （H.brasiliensis ），MnASI1（M.notabilis 1），MnASI2（M.notabilis 2）Fig.3　Comparison of 7 ASI amino acid sequencesThe known structural features of ASI are indicated.Bidirectional arrows are Kunitz -type protein common sequences.Round -headed arrows are disulfide bonds.Inverted triangle marks the catalytic residue against Subtilisin ， followed by the reactive loop in the box.Six -pointed star marks the alpha -amylase binding key SI （L.chuanxiong ），BASI （H.vulgare ），RASI （O.sativa Japonica Group ），WASI （T.aestivum ），HbASI （H.brasiliensis AJG01567.1），MnASI1（M.notabilis1 EXB74706.1），MnASI2（M.notabilis 2）第 61 卷四川大学学报（自然科学版）第 2 期图5　33条ASI 序列保守基序分析Fig.5　Conserved motifs analysis of 33 ASI sequences图4　LASI 与（a ）单子叶植物、（b ）双子叶植物及（c ）所有植物来源ASI 序列变异分析Fig.4　Shinon protein variability of LASI with （a ） monocotyledonous plants ， （b ） dicotyledonous plants and （c ） all plants第 2 期黄斌翰，等： ASI 基因家族结构及表达分析第 61 卷RASI 主要在水稻叶鞘组织中表达.在胁迫处理下，叶鞘组织中的RASI 表达均出现显著下调，在叶片组织中均发生上调，在根中既出现上调（干旱胁迫处理下）也存在下调（盐胁迫处理下），提示RASI 在水稻不同组织中可能具有不同的生物学功能.4　讨论本文通过生物信息学方法鉴定出33条不同植物来源ASI 基因，它们编码蛋白质的平均分子量为22 kD 左右，这与Kunitz 型蛋白酶抑制剂家族的分子量大小吻合［15］.并且，所有ASI 成员的N -末端均含有Kunitz 型蛋白酶抑制剂家族的特征序列（［LIVM ］-x -D -x -［EDNTY ］-［DG ］-［RKHDENQ ］-x -［LIVM ］-（x ）5-Y -x -［LIVM ］）［16］，表明ASI 属于Kunitz 家族成员，然而ASI 的进化地位，尚待进一步研究.信号肽分析显示大部分ASI 蛋白带有亲水性的信号肽，提示ASI 可能为分泌蛋白，这一结果与亚细胞定位预测一致，即ASI 可能定位于质外体或细胞壁中.质外体是位于细胞膜与细胞壁之间的区域，该区域含有大量的保护酶与活性小分子，对植物发挥保护性作用［17， 18］，由此推测ASI可能具有保护植物的功能.系统发育分析显示基于ASI 的系统进化树与基于APG Ⅳ系统构建的物种分类基本一致，表明ASI 基因进化比较保守，可以为物种分类研究提供一定的参考依据.在所分析的序列中，有7种植物报道了2~3条ASI 并系同源基因.其中来源于黄花蒿、川桑以及稷的并系同源基因基本按物种关系聚类；而来源于毛果杨、牧豆、野大豆与寡头二蕊草的并系同源基因没有完全按照物种聚类，推测在这些物种中ASI 基因有的拷贝分化时间较早，变异积累较多，功能产生差异，因此在进化树中有可能按照基因功能聚类.ASI 的结构体现出较强的保守性，例如Kunitz 特征序列均位于基序1，抑制枯草杆菌蛋白酶的关键位点及区域分布于基序2，而发挥α-淀粉酶抑制活性的区域均位于基序4.然而，ASI 部分区域的氨基酸序列仍然体现较高的变异性，例如，ASI 含有抑制枯草杆菌蛋白酶和α-淀粉酶的两个位点，分图6　ASI 基因顺式作用元件分析Fig.6　Characterization of cis -acting elements of the ASIgenes图7　3种处理条件下水稻RASI 基因表达情况Fig.7　RASI gene expression under free stress ， droughtstress and salt stress第 61 卷四川大学学报（自然科学版）第 2 期别位于N-末端（例如BASI中的Thr110残基，其通过氢键和分子内作用力稳定与枯草杆菌蛋白酶结合的活性环构象以维持抑制活性［5］）和C-端（例如BASI中的Glu190残基，其通过水分子与Ca2+形成氢键，参与结合α-淀粉酶从而发挥抑制作用［2， 19］）.比较两个位置的香农熵值，发现Thr110的保守性更强，香农熵值仅有1.014，而Glu190的变化较大，香农熵值为2.883，推测Thr110存在较大的选择压力.与此同时，双子叶植物ASI的基序6中，含有一段变异程度较高（平均熵值2.14）的20肽序列（LASI 的Gly137-Asn156），然而，单子叶植物ASI的对应序列的变异程度却较低，平均香农熵值为1.21，尽管导致该现象的原因尚不清楚，但该区域有可能作为区分单双子叶植物ASI的一个重要标志.分析ASI基因的启动子区域，共鉴定到光、脱落酸和茉莉酸甲酯响应元件等10余种响应生物及非生物胁迫的顺式作用元件，提示ASI可能在植物生长发育及胁迫响应过程中发挥作用.以上分析结果已在不同植物中获得验证，例如大麦、水稻、小麦、橡胶树、川桑、川芎等［8， 9， 20-23］.同时，本文还发现RASI基因的组织表达模式较复杂，其主要分布在幼苗期的水稻叶鞘.此前已有研究发现在翅豆筛管中大量存在一种Kunitz蛋白酶抑制剂，其在防御系统中可能发挥作用，以抵御吸食韧皮部汁液的昆虫或在受伤时入侵的病菌［24］.同时，也有研究报道Kunitz型蛋白酶抑制剂基因在麻风树的根和茎中表达较高，以应答害虫对根、茎部的入侵［25］.水稻叶鞘系维管系统组分，其韧皮部由筛管分子、伴胞和薄壁细胞组成［26］，同时水稻叶鞘中营养物质丰富，易受到昆虫如褐飞虱的吸食，而病菌又易通过伤口侵入水稻，导致叶鞘腐败病等［27］.RASI在叶鞘中的高表达，可能有助于减轻病虫害对水稻的伤害.水稻幼苗受到干旱与盐胁迫后，RASI基因在不同组织中的表达变化呈现多样化，其中在叶中的表达出现明显上调，表明RASI可能参与了对两种胁迫的响应过程，这一结果与RASI 启动子区域中含有ABA响应元件相吻合；但叶鞘中RASI在胁迫后表达下调，推测叶鞘不是RASI 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化学分析计量CHEMICAL ANALYSIS AND METERAGE第30卷，第6期2021年6月V ol. 30，No. 6Jun. 202144doi ：10.3969／j.issn.1008–6145.2021.06.010碳酸氢铵–二乙三胺五乙酸浸提–电感耦合等离子体发射光谱法测定碱性土壤中交换性钠杨娜，张文舒，朱丽，郝冠军（上海市园林科学规划研究院，上海　200230）摘要　建立一种使用碳酸氢铵–二乙三胺五乙酸（AB–DTPA ）浸提液提取–电感耦合等离子体发射光谱（ICP–OES ）法快速测定土壤中交换性钠的方法。

优化的样品处理条件：称取5 g 土壤样品，加入AB–DTPA 浸提液75 mL ，在25 ℃下以170 r ／min 振荡浸提15 min ，用ICP–OES 法测定提取液中的交换性钠。

结果表明，钠离子的质量浓度在0～10.0 mg ／L 范围内与发射光谱强度具有良好的线性关系，线性相关系数r 2=0.999 9，检出限为0.010 cmol ／kg 。

用该方法测定标准物质GBW 07413a 和GBW 0714a ，测定结果的相对标准偏差分别为6.25%、7.04%（n =6），加标回收率为93.3%～103.3%。

AB–DTPA 浸提法测定结果与乙酸铵浸提法无明显差异。

该方法适合于碱性土壤交换性钠的检测。

关键词　碳酸氢铵–二乙三胺五乙酸浸提液；电感耦合等离子体发射光谱法；碱性土壤；交换性钠中图分类号：O657.7 文献标识码：A 文章编号：1008–6145(2021)06–0044–04Determination of exchangeable sodium in alkaline soilby ammonium bicarbonate–diethylenetriamine pentaacetic acid extraction–ICP–OESYang Na ，Zhang Wenshu ，Zhu Li ，Hao Guanjun(Shanghai Academy of Landscape Architecture Science and Planning ，Shanghai 200230，China)Abstract A method for fast determination of exchangeable sodium in alkaline soil by using ammonium bicarbonate –diethylenetriamine pentaacetic acid(AB–DTPA) extraction and inductively coupled plasma optical emission spectrometry(ICP–OES) was established. The optimized sample processing conditions: 5 g of soil samples were weighed and 75 mL of AB–DTPA extraction was added to extract at 25 ℃ under 170 r ／min oscillation for 15 min ，then the exchangable sodium in the extract was determined by ICP–OES. The mass concentration of sodium ion had good linear relationship with the intensity of emission spectrum in the range of 0–10.0 mg ／L ，the linear correlation coef ficient r 2=0.999 9，the detection limit was 0.010 cmol ／kg. The relative standard deviations of determination results were 6.25%，7.04%(n =6)，the recovery of standard addition was 93.3%–103.3%. There was no signi ficant difference between AB–DTPA extraction method and ammonium acetate extraction method. The method is suitable for the determination of exchangeable sodium in alkaline soil.Keywords ammonium bicarbonate–diethylenetriamine pentaacetic acid; inductively coupled plasma atomic emission spectrometry; alkaline soil; exchangeable sodium土壤养分是土壤肥力的重要标志，阳离子交换量和交换性盐基离子作为土壤养分的重要指标，在一定程度上反映了土壤肥力［1］，自盖德罗依茨创造碱土发生的物理化学理论以来，交换性钠一直被认为是土壤碱化过程的本质［2］。

Vol.41No.4Apr.2021上海交通大学学报（医学版）JOURNAL OF SHANGHAI JIAO TONG UNIVERSITY (MEDICAL SCIENCE)USP47通过促进抗凋亡蛋白表达介导头颈鳞癌细胞顺铂耐药童桐，秦星，谢非，蒋英英，石剑波，张建军上海交通大学医学院附属第九人民医院·口腔医学院口腔颌面头颈-肿瘤科，国家口腔疾病临床医学研究中心，上海市口腔医学重点实验室，上海市口腔医学研究所，上海200011［摘要］目的·探讨去泛素化酶47（ubiquitin carboxyl-terminal hydrolase 47，USP47）对头颈鳞癌细胞顺铂耐药的影响及潜在机制。

方法·通过梯度增加顺铂浓度，构建头颈鳞癌顺铂耐药细胞株；通过实时定量PCR 和蛋白免疫印迹实验检测耐药株中USP47的表达情况；利用慢病毒载体构建USP47基因过表达或敲除的稳转细胞株，MTT 法检测USP47表达升高或降低对头颈鳞癌细胞顺铂耐药性的影响；并检测USP47对癌细胞增殖、克隆形成和凋亡等生物学行为的影响；通过免疫印迹实验检测USP47对抗凋亡蛋白X-连锁凋亡抑制蛋白（X-linked inhibitor of apoptosis protein ，XIAP ）和重组人B 细胞淋巴瘤因子2xL （recombinant human B-cell leukemia/lymphoma 2xL ，Bcl-xL ）的表达及泛素化水平的影响。

结果·USP47在头颈鳞癌顺铂耐药细胞株中显著高表达；过表达USP47后，顺铂对头颈鳞癌细胞的半数抑制浓度（half maximal inhibitory concentration ，IC 50）明显升高，而敲除USP47可以降低头颈鳞癌顺铂耐药细胞株的IC 50；过表达USP47的头颈鳞癌细胞，其增殖和克隆形成能力均被抑制，而抗凋亡能力明显增强。

植物学通报 2005, 22 (6): 715￣722 Chinese Bulletin of Botany专题介绍植物 SNARE 蛋白的结构与功能①鲍永美 王州飞 张红生 ②(南京农业大学作物遗传与种质创新国家重点实验室 南京 210095)摘要在真核生物细胞囊泡运输过程中的膜融合主要是由SNARE蛋白介导的, SNARE蛋白的结构高度保守。

研究发现, 植物中的SNARE蛋白促进植物细胞板形成, 能与离子通道蛋白相互作用, 有利于 植物的正常生长发育, 能提高植物的抗病性及参与植物的向重力性作用。

应用基因组学和蛋白质组学 技术结合细胞学水平上的分析方法有助于深入揭示植物SNARE蛋白家族成员的功能, 明确SNARE蛋 白在信号转导途径中的作用, 阐明动植物免疫系统的区别和联系。

关键词 SNARE蛋白, 结构, 功能Structure and Function of SNAREs in PlantBAO Yong-Mei WANG Zhou-Fei ZHANG Hong-Sheng②(State Key Laboratory of Crop Genetics and Germplasm Enhancement, Nanjing Agricultural University, Nanjing 210095)AbstractThe membrane fusion in vesicle trafficking in the cells of eukaryotic organisms ismediated by soluble-N-ethyl-maleimide-sensitive fusion protein attachment protein receptor (SNARE) proteins, which are highly conserved in their structures from various species, including yeast, animals and plants. Increasing research has demonstrated many tissue- or subcellularspecific components of SNAREs involved in the formation of the cell plate, interacting with ion channel proteins, and gravity sensing in plants. SNARE proteins might play important roles in plant growth, development and response to abiotic and biotic stresses. The application of genomics and proteomics approaches, as well cytological methods, will accelerate our understanding of diverse functions of the plant SNARE family and their specific location in the signal transduction pathway, and the differentiation and relation between animal and plant immunity systems. Key words SNARE protein, Structure, Function 植物细胞内可溶性物质的运输主要是通 过细胞内的囊泡(vesicles)在不同细胞器膜结 构间穿梭运输而实现(Sanderfoot et al., 1999), 囊泡充当了整个系统的运输工具。