Abscisic Acid- and Low Temperature-increased Gene Expression along with Enhancement of Freezing Tolerance in Physcomitrella patens
Manabu Nagao, Anzu Minami, Keita Arakawa, Seizo Fujikawa* and Daisuke Takezawa
Institute of Low Temperature Science; *Graduate School of Agriculture, Hokkaido University, Sapporo, Japan
Cold acclimation at low, non-freezing temperatures enhances ability of plants to tolerate freezing at various subzero temperatures, accompanying physiological and morphological alterations at the cellular level. Abscisic acid (ABA) is postulated to be crucial to the development of freezing tolerance during the cold acclimation process in plants. A number of genes are reported to be induced by cold acclimation and ABA treatment in higher plants, and the proteins encoded by these genes are thought to play a role in protecting plant cells from freezing injury. However, specific functions of these cold- and ABA-induced genes in vivo are not well understood because of lack of studies on genetic mutants deficient in each gene. In the present study, we attempted to use Physcomitrella patens, in which specific genes can be disrupted by homologous recombination,
as a model system to identify the roles of the cold- and ABA-induced genes and to clarify the basic mechanism of freezing tolerance in plants. Equilibrium freezing test of P. patens protonema cells revealed that the cells grown under the normal growth condition are labile to freezing. ABA treatment, however, dramatically increased the freezing tolerance within several hours. Cold treatment at 4 淸 also increased the freezing tolerance, but the increase was far less than that achieved by ABA. We examined ultrastructures of freeze-labile and tolerant protonema cells by freeze-fracture electron microscopy. When non-treated control cells were subjected to freezing at –4 淸, ?aparticulate domains? and ?fracture jump lesions?, structures characteristic to freeze-injured cells, were observed in the plasma membrane. However, these structures were not observed in the ABA-treated cells frozen at the same temperature. Changes in gene expression during ABA and cold treatment were investigated by Northern analysis using probes prepared from clones isolated by differential display and EST clones with similarity to known stress-induced genes. The results indicated that the majority of the ABA-increased genes were also responsive to low temperature. Possible roles of these genes in the
development of freezing tolerance in the moss protonema cells will be discussed in the session.
Phylogenetic analysis of land plants using introns within the nad7 gene
Misa Nakamura, Yukihiro Kabeya, Kazuyoshi Hashimoto, Naoki sato Department of Molecular Biology, Faculty of Science, Saitama University
Urawa, Saitama Prefecture, 338-8570
Bryophytes represent the earliest lineage of land plants. However, many problems of phylogenetic relationships within the bryophytes and with the tracheophytes remain. Are bryophytes monophyletic or paraphyletic? Which of the three groups of bryophytes (mosses, liverworts and hornworts) is the closest lineage to chlorophytes and tracheophytes? Recent analyses often report that hornworts are the first land plants and mosses plus liverworts make a sistergroup to tracheophytes. The nad7gene in the mitochondrial genome encodes the subunit 7 of the NADH dehydrogenase complex I. In most of the seed plants, the nad7gene is encoded in the mitochondrial genome, while it
is encoded in the nuclear genome in fungi and animals. The nad7gene contains four group II introns in many angiosperms. They are called intron 1, 2, 3 and 4, respectively. Hashimoto and Sato (2001) reported that the moss Physcomitrella patens also have two group II introns exactly corresponding to the intron 1 and 2 of angiosperms, and suggested that this information potentially useful in phylogenetic analysis. On the other hand, in liverworts, Marchantia polymorpha, the nad7gene is encoded in the nuclear genome. Another copy exists in the mitochondrial genome as a pseudogene, which is interrupted by six stop codons. This pseudogene contains two group II introns at the positions different from those of angisperm introns. We used the presence of four mitochondrial introns of the nad7gene as a phylogenetic marker to estimate the phylogenetic relationships between land plants. Total DNA was isolated from various plant materials according to the CTAB method or extraction with phenol and chloroform, then purified by CsCl density gradient centrifugation. For small amounts of material, DNA was isolated by DNeasy Plant Maxi/Mini Kit (QIAGEN). Each intron segment was amplified by PCR using the primers corresponding to the flanking exon sequences. The presence or absence and the length of the four introns as well
as the total length of the nad7 gene were analyzed. No intron was detected in the nad7 gene of chlorophytes and hornworts. They were not present in liverworts either, However, another intron is estimated to be present in both Jungermanniales and Marchantiales. Mosses had introns 1 and 2. Intron 3 first appeared in ferns. In some species, the lack of intron 2 and the presence of another intron could be considered. Intron 4 first appeared in several species of gymnosperms. Each intron (intron 1, 2, 3 or 4) was conserved well in most angiosperms. However, intron 3 was lost in Calystegia japonica and Perilla frutescens (See the table below). These results suggest that hornworts are the earliest land plants. plant materials (number of species)
Intron 1 Intron 2 Intron 3 Intron 4 Unidentified intron angiosperms (15)
+ + + - + - gymnosperms
(3)
+ + + - + - - ferns
(10)
+ + - + - - - mosses
(3)
+ + - - - liverworts
(7)
- - - - + hornworts
(2)
- - - - - chlorophytes
(4) - - - - -
Physiological function of a germin-like protein with Mn-SOD activity from a moss, Barbula unguiculata
M. Nakata, Y. Watanabe, T. Shiono, T. Suzuki, M. Matsuzaki, I. Yamamoto, and T. Satoh
Deptartment of Biological Science, Graduate School of Science, Hiroshima University, Higashi-Hiroshima 739-8526, Japan
We isolated an extracellular protein with Mn-SOD activity from a moss, Barbula unguiculata(1). The protein was identified as one of germin-like proteins (BuGLP) from the sequence of the cDNA. Although germin from wheat has oxalate oxidase activity, BuGLP has no its activity. Our purpose in this study is to bring out physiological functions of BuGLP. First, the relations between cell growth and expression of BuGLP were investigated. Although BuGLP mRNA accumulated more in the exponential phase of growth, BuGLP protein accumulated more in the later phase. Then, effects of salt stress on BuGLP expression was studied. Salt stress induced the expression of BuGLP at the transcriptional level (Fig. 1B) and accumulation of the protein level (Fig. 1C). Most of BuGLP protein, however, was eluted in the medium, suggesting that BuGLP binds with a weak force to the cell wall. To examine relations between amount of superoxide and
BuGLP expression, effect of methyl viologen was determined. Although methyl viologen suppressed accumulation of BuGLP mRNA, amount of BuGLP protein was not affected. Since the Mn-SOD was conceivable to participate in the formation of lignin, phenolic compounds which appear to constitute lignin were determined. They were detected in the cell wall fractions of Barbula, Physcomitrella and Marchantia. The physiological functions of BuGLP is discussed (Figure 2). Key words : extracellular Mn-SOD, germin-like protein, salt stress protein, cell wall, moss
Reference
1. T. Yamahara, T. Shiono, T. Suzuki, K. Tanaka, S. Takio, K. Sato, S. Yamazaki, and T. Satoh (1999) Isolation of germin-like protein with manganese superoxide dismutase activity from cells of a moss, Barbula unguiculata. J. Biol. Chem.47: 33274-33278
A molecular phylogenic study of the Sematophyllaceae (Hypnales, Musci) and their allies
Hiromi Tsubota1, Hiroyuki Akiyama2, Tomio Yamaguchi1 and Hironori Deguchi1
1Department of Biological Science, Graduate School of Science, Hiroshima University, Japan, 2Museum of Nature and Human Activities, Hyogo, Japan
Circumscription of the Sematophyllaceae is controversial, and the taxonomic position of Brotherella, Pylaisiadelpha, Heterophyllium, Trismegis tia, Wijkia and their allies have been repeatedly discussed. Phylogenetic relationships between the Sematophyllaceae and the Hypnaceae have been the focus of much attention in recent years (e.g., Tsubota et al. 1999, Arikawa & Higuchi 1999, De Luna et al. 2000, Tsubota et al. 2000 and Tsubota et al. 2001). Phylogenetic analyses were carried out to investigate the phylogenetic position of Brotherella and its allies and the relationships between the Sematophyllaceae and the Hypnaceae, based on the chloroplast ribulose 1,5-bisphosphate carboxylase/oxygenase large subunit (rbc L) gene sequences. Here phylogenetic inferences were presented based on the log-likelihood difference of alternative trees constructed by the neighbor-joining (NJ), minimum-evolution (ME), maximum-parsimony (MP) and maximum-likelihood (ML) methods. This study suggests that (1) the Sematophyllaceae (s. lat.) are a monophyletic entity; (2) the Hypnaceae are not monophyletic,
with no particular clade accommodating members of the family; (3) the genera Wijkia, Trismegistia and Acanthorrhynchium do not form a single clade; and (4) the genera Brotherella, Pylaisiadelpha, Heterophyllium, Wijkia, Isocladiella, and some species treated as members of the Hypnaceae, such as Hypnum tristo-viride and Isopterygium tenerum, should be placed in the Sematophyllaceae (s. lat.). The present research shows that the Sematophyllaceae both sensu Seki (1968) and sensu Heden郭 & Buck (1999) are not monophyletic. Since the monophyly of the Sematophyllaceae (s. lat.) including Brotherella and its allies is well supported, we propose that the Sematophyllaceae should be treated in a wider sense. This suggests the necessity of the reconsideration of familial circumscription within the order Hypnales. The Entodontaceae (s. str.) and the Thuidiaceae (s. str.) have been resolved as monophyletic groups. The Thuidiaceae (s. str.), one of the groups with a single costa, should be placed near the Sematophyllaceae (s. lat.) together with the Entodontaceae (s. str.). These results mean that a single costa is an ancestral character state with multiple transitions to a double costa state, and do not concur with the systems of Vitt (1984) and Buck & Vitt (1986), where it was suggested that the groups with a single costa
were divergent from the hypnalean ancestor at an early evolutional stage and followed by groups with a double costa.
Monoplastidic meiosis in Marchantiales
*Masaki Shimamura, Akane Kitamura and Hironori Deguchi
(*Contact person) Department of Biological Science, Graduate School of Science, Hiroshima University, Kagamiyama 1-3-1, Higashi-hiroshima-shi, Hiroshima 739-8526, Japan
Sporogenous tissues of Marchantiales were examined for the occurrence of monoplastidic meiosis. The early sporocyte of Wiesnerella denudata and Marchantia polymorpha contained a single large plastid. Plastid division occured in advance of nuclear division and the four resultant plastids migrate to the future spore domains. The four daughter plastids continued to divide before nuclear division. Positioning of plastids seem to establish the meiotic division planes before nuclear division. Such morphogenetic plastid division and migration have been considered as symplesiomorphic character among all bryophyte clades, lycopsid pteridophytes, and the coleochaetalean algae (Brown & Lemmon 1997). However, in
liverworts, polyplastidic meiosis is common and monoplastidic meiosis had been observed in only four species belonging to Marchantiales, Metzgeriales, Monocleales and Haplomitriales (Renzaglia et al. 1994, Shimamura et al. 2000). This paper reports two additional species with monoplastidic meiosis for liverworts: Wiesnerella denudata and Marchantia polymorpha(Marchantiales). These facts could be explained by postulating that the divergence of liverworts groups (orders) occurred rapidly in the ancient radiation of land plant. Thus plesiomormorphic characters could have remained in a mosaic pattern with polyplastidic meiosis having been established independently in various lineages of liverworts. Absence of cytoplasmic lobing of the sporocyte in Marchantiidae has been accepted as a decisive character distinguishing Marchantiidae from Jungermanniidae (Schuster 1983). However, slightly lobed sporocytes can be observed in Monoclea(Renzaglia et al. 1994) and Dumortiera (Shimamura et al. 2000) of Marchantiidae. Wiesnerella and Marchantia have sporocyte similar to that of Monoclea and Dumortiera having a single plastid, which becomes more than four by subsequent plastid division before nuclear division. In general, monoplastidy of
lower land plant cells is maintained through meiosis and newly formed spores have a single plastid (Brown & Lemmon 1997). On the other hand, the sporocyte shape of Wiesnerella and Marchantia are also similar to that of other Marchantiidae species with polyplastidic meiosis (e.g.Conocephalum, Reboulia, Monosoleniu) in that cytoplasmic lobing does not occur before nuclear division. The characters in Wiesnerella-Marchantia type meiosis seems to be transitional between that of Monoclea-Dumortiera type and other Marchantiidae type.
A GENOMICS BASED APPROACH TO FIND GENES INVOLVED IN ASYMMETRIC CELL DIVISION IN THE MOSS PHYSCOMITRELLA PATENS
Tomomichi Fujita1, Motoaki Seki2, Asako Kamiya2, Ikuo Uchiyama1, Piero Carninci2, Yoshihide Hayashizaki2, Kazuo Shinozaki2, and Mitsuyasu Hasebe1
1NIBB, Okazaki, Japan, 2RIKEN, Tsukuba, Japan
Asymmetric cell division is of fundamental importance in the generation of the overall cellular pattern in plants. Its molecular mechanism, however, remains unknown. The
moss Physcomitrella patens will be a good system to dissect this molecular mechanism, not only because its body plan is relatively simple, making a study at a single cell level feasible, but also because it is the only plant in which gene targeting exhibits a high rate of success. We are currently trying to develop a comprehensive system to screen mutants affecting the asymmetric cell division during the regeneration step of protoplasts in P. patens.
We first constructed a full-length cDNA library based on 5' cap trapper method from protonemal cells of P. patens and determined the sequences from 5' and 3' ends of multiple cDNA clones. The sequencing data was clustered and annotated by BLAST search. Sequence analyses of the first 900 cDNAs revealed that more than 91% of the clones appeared to represent full-length clones having the first, putative start codon and that about 41% of the clones were no matches to any sequences in the database. We will also construct full-length cDNA libraries from auxin or cytokinin treated protonemal cells and determine the sequences from both ends of 15,000 clones, respectively. These will serve as good sources of EST database and for functional analysis of gene products. Individual full-length cDNAs then are subcloned under a
constitutive promoter and transiently expressed in moss protoplasts to test the effect of each clone on the regeneration step of protoplasts. The regeneration step includes several interesting aspects such as cell polarity, asymmetric cell division, or cell differentiation. We will present the results of sequencing of the full-length cDNAs and the progress of this project.
Identification and characterization of API1 gene encoding a kinesin-like protein, which is predominantly expressed in apical cells in the moss, Physcomitrella patens.
Yuji HIWATASHI 1, 2, Tomoaki NISHIYAMA 2, Takashi MURATA 2, Tomomichi FUJITA 2, and Mitsuyasu HASEBE 1, 2, 3,
1Dept. Mol. Biomechanics, Grad. Univ. Advanced Studies, Okazaki 444-8585, 2Spec. Mechanisms 2, Natl. Inst. Basic Biol., Okazaki 444-8585
During the course of moss development, an apical cell produces a multicellular protonemata and gametophores with different specialized cells. To form plant body of mosses requires cell divisions in which one daughter cell is kept to be an apical cell
and the other cell differentiate to another cell type. This type of asymmetric cell division is similar to that of stem cells in animals. For the first step to analyze the molecular mechanism of asymmetric cell division of an apical cell, gene-trap and enhancer-trap systems in the moss Physcomitrella patens were established. Several lines with apical cell specific expression were isolated. Of them, one gene-trap line, Apicar1, showing a predominant expression of uidA reporter gene at the apical cells of caulonemata, rhizoids, and gametophores, was characterized. UidA-fused fragments were amplified by 5?-RACE method using uidA-specific primers, and its expected full-length cDNA was determined by both 3? and 5? RACE methods. Sequence analysis of the isolated cDNA revealed that uidA gene was fused to the 5? untranslated region of a kinesin-like gene, named API1(Api car1). According to a phylogenetic analysis of API1 and other members of kinesin gene family, API1gene formed a new subfamily of kinesin-like protein with one of Arabidopsis kinesin-like genes. Kinesin-like genes are classified into several groups, and each group has its specific function. This suggests that API1 may have a novel function different from other kinesin-like proteins in other subfamilies. To visualize intracellular localization of API1 protein,
the GFP gene was inserted just before the stop codon of API1gene using homologous recombination. The API1-GFP fusion protein was predominantly localized at a forming cell plate in dividing caulonemal cells, suggesting that API1 protein is related to cell plate formation during cytokinesis. In a non-dividing caulonemal cell, the API1-GFP fusion protein was detected in a nucleus and a distal part of cytosol in an apical cell, although their function is not yet revealed. API1D C-GFP gene, coding the GFP protein fused after the motor domain of API1 lacking its C-terminus, was transformed. The truncated API1 protein was not localized in its original places. This indicates that the C-terminal region adjacent to motor domain requires for nuclear and cell plate localization. To clarify the function of API1 in the asymmetric cell division of the apical cell, API1gene was disrupted by the gene-targeting technique. Stable transgenic lines constitutively expressing full-length or truncated API1 protein were generated. The phenotype of these transgenic lines will be reported and discussed.
Cloning of Nuclear-Encoded T7 Phage-Type RNA Polymerase Genes from Physcomitrella patens
Yukihiro Kabeya, Kazuyoshi Hashimoto, Naoki Sato
Department of Molecular Biology, Faculty of Science, Saitama University,
Urawa, Saitama Prefecture, 338-8570
Plastids and mitochondria employ different strategies for expressing their genetic information. In plastids, both nuclear-encoded phage-type RNA polymerase ( RPOT ) and plastid-encoded eubacteria-type RNA polymerase are known to function. In contrast, only RPOT is present in mitochondria, whereas RPOT does not exist in cyanobacteria, which is considered to be related to the origin of plastids. It is thought that plastid type RPOT gene was produced by the duplication of the mitochondria-type RPOT gene. Currently, the presence of the plastid type RPOT gene is not known in green algae. To shed light on the evolution of plastid type RPOT gene, we isolated RPOT genes from the moss Physcomitrella patens. We used the polymerase chain reaction ( PCR ) strategy to clone the P. patens RPOT genes. To amplify portions of RPOT genes that might be present, PCR was carried out with genomic DNA using degenerate primers, the design of which was based on amino acid sequence blocks conserved among higher plant RPOT sequences. We
obtained a short fragment of RPOT gene as a result of PCR. In addition, the partial sequence of another RPOT gene was registered in the EST database. These portions of RPOT genes were used as probes to screen a cDNA library constructed from protonematal RNA. We isolated two cDNA clones for RPOT in P. patens. They were named PpRPOT1 and PpRPOT2, respectively. These genes encode putative polypeptides of 1087 and 1065 residues, respectively. Putative transit sequence was found in each of the putative gene products. To examine the subcellular localization of the PpRPOT1 and PpRPOT2, fusion constructs containing each of the putative N-terminal transit peptides and the GFP coding region were prepared. These constructs were introduced into the moss protoplasts by polyethylene glycol-mediated transformation. One day after transformation, the protoplasts were examined by fluorescence microscopy. These constructs gave rise to GFP fluorescence in typical mitochondrial structures of punctate morphology. It seems that RPOT1 and RPOT2 are both mitochondrial RNA polymerases. However, we did not find a plastid type RPOT in P. patens. To surmise when the plastid type RPOT gene has been generated, maximum-likelihood ( ML ) trees were generated by the PROTML program using an alignment of RPOTs and T7 phage RNA polymerase
that was assembled using the CLUSTAL X program. We also constructed a neighbor-joining tree and both were consistent. As a result of phylogenetic analysis, the two RPOTs of P. patens formed a sister group to the counterparts in higher plants. The P. patens RPOT gene had branched before the plastid type RPOT gene had been generated by gene duplication. These results suggest that the gene duplication of RPOT geneis a rather recent event during the evolution of land plant.
信号转导 061M5007H 学期：2015-2016学年秋| 课程属性：| 任课教师：谢旗等 教学目的、要求 本课程为细胞生物学专业研究生的专业基础课，同时也可作为相关专业研究生的选修课。细胞信号转导是细胞生物学学科进展最快的研究领域之一，信号转导的概念已经开始深入到生命科学的各个领域。本课程内容涵盖动植物受体、G蛋白、环核苷酸第二信使、质膜磷脂代谢产物胞内信使、酶活性受体、蛋白质可逆磷酸化、泛素蛋白化及其对基因表达的调控、信号转导途径的多样性、网络化和专一性等方面的研究现状和进展。 预修课程 生物化学、分子生物学 教材 生命科学学院 主要内容 第一章绪论（3学时，教师：谢旗）细胞信号转导的研究对象和研究意义，细胞信号的主要种类，细胞化学信号分子与信号传递途径的特征。真核生物的蛋白激酶，蛋白磷酸酶，蛋白质可逆磷酸化对信号转导的调节方式，蛋白质可逆磷酸化与基因表达调控，蛋白质可逆磷酸化在细胞信号中的意义。蛋白质稳定性与信号转导。第二章植物免疫的表观遗传调控（3学时，教师：郭惠珊）表观遗传调控包含RNA干扰、DNA修饰、组蛋白翻译后修饰和染色质重塑等各种过程互相交叠，共同调控基因组表观修饰的动态平衡；除了影响生长和发育，表观遗传调控的另一重要功能是抗病免疫作用。本讲将着重介绍植物表观遗传途径及其抗病免疫信号的调控作用。第三章MicroRNA介导的信号（3学时，教师：郭惠珊）microRNA 广泛存在于生物体内，是生物体保守机制RNA沉默过程产生并具有序列特异性调控功能的一类非编码小分子RNA。本课程主要讲授植物microRNA的产生、加工、特性及其调控作用的基本生物学过程；以及植物miRNAs和其他小分子RNA参与植物生长素信号途径和其他植物生理性状的调控作用。第四章钙离子通道及信号转导（3学时，教师：陈宇航）钙离子是生命活动的必需元素，基本分布和内稳，代谢平衡和疾病；钙离子发挥重要生物学功能，简述历史发现，作为第二信使的化学基础，功能调控的基本模式，以钙结合蛋白为例子展开介绍钙离子发挥功能调控的分子结构基础等；介绍钙离子信号转导系统的组成，
细胞信号转导 李婧 2015212351 一、名词解释 内分泌 接触依赖性通讯 受体 G蛋白 第二信使 二、单项选择题 1、下列不属于信号分子的是 A.K+ B.cAMP C. cGMP D.Ca2+ 2、下列那个不是信号转导系统的主要特性 A.特异性 B.放大效应 C.整合作用 D.传递作用 3、（）是细胞表面受体中最大的多样性家族 A.G蛋白偶联受体 B.RTK C.Notch D.细胞因子 4、G蛋白偶联受体中（）是分子开关蛋白 A.G α B.Gβ C.GΘ D.Gγ 5、G蛋白偶联的光敏感受体的活化诱发（）的关闭 A.cAMP–PKA信号通路 B.Notch信号通路 C.JAK-STAT信号通路 D. cGMP门控阳离子通道 6、（）信号对细胞内糖原代谢起关键调控作用 -Ca2+ B.DAG-PKC C. cAMP–PKA D.RTK-Ras A.IP 3 7、（）的主要功能是引发贮存在内质网中的Ca2+转移到细胞质基质中，使 胞质中游离Ca2+浓度提高 B.PIP2 C.DAG D.PKC A. IP 3 8、（）主要功能是控制细胞生长、分化，而不是调控细胞中间代谢 A.RTK B. PKC C.PKB D.Wnt 9、Hedgehog信号通路控制 A.糖原代谢 B.细胞凋亡 C.细胞分化 D.氨基酸代谢 10、细胞通过配体依赖性的受体介导的（）减少细胞表面可利用受体数目。 A. 抑制性蛋白产生 B.内吞作用 C.敏感性下调 D.消化降解 三、多项选择题 1、细胞分泌化学信号可长距离或短距离发挥作用，其作用方式分为 A.内分泌 B.旁分泌 C.通过化学突出传递神经信号 D.外分泌 2、下列（）是糖脂 A.霍乱毒素受体 B.百日咳的毒素受体 C.甲状腺受体 D.胰岛素受体 3、下面关于受体酪氨酸激酶的说法正确的是 A.是一种生长因子类受体 B.受体蛋白只有一次跨膜 C.与配体结合后两个受体相互靠近，相互激活 D.具有SH2结构域
植物生长发育的各个阶段, 包括胚胎发生、种子萌发、营养生长、果实成熟、叶片衰老等都受到多种植物激素信号的控制。人们对植物激素的生物合成途径、生理作用已有大量阐述，在生产上的应用也已取得很大进展，但对其信号转导途径的认识并不是很全面。今天小编和大家聊一聊，9大类植物激素信号转导途径。 1.生长素 与生长素信号转导相关的三类蛋白组分是:生长素受体相关SCF复合体(SKP1, Cullin and F-box complex)、发挥御制功能的生长素蛋白(Aux/IAA)和生长素响应因子(ARF)。早期响应基因有Aux/IAA基因家族、GH1、GH3、GH2/4、SAUR基因家族、ACS、GST。生长素信号转导通路主要有4条: TIR1/AFBAux/IAA/TPL-ARFs途径、T MK1-IAA32/34-ARFs途径、TMK1/ABP1-ROP2/6-PINs或RICs 途径和SKP2AE2FC/DPB途径。 2.细胞分裂素
细胞分裂素信号转导途径是基于双元信号系统（TCS），通过磷酸基团在主要组分之间的连续传递而实现。双元信号系统主要包含3类蛋白成员及4次磷酸化事件: (ⅰ)位于内质网膜或细胞膜的组氨酸受体激酶(histidine kinases, HKs)感知细胞分裂素后发生组氨酸的自磷酸化；(ⅱ)将组氨酸残基的磷酸基团转移至自身接受区的天冬氨酸残基上；(ⅲ)受体天冬氨酸残基上的磷酸基团转移至细胞质的组氨酸磷酸化转移蛋白(His-containing phosphotransfer protein, HPs)的组氨酸残基上；(ⅳ)磷酸化的组氨酸转移蛋白进入细胞核并将磷酸基团转移至A类或B类响应调节因子(response regulators, ARR s)。在拟南芥中已知的细胞分裂素受体有AHK2、AHK3和AHK4 3个，AHP有6个(AHP1?6)，A类和B类ARR分別有10个和1 2个，它们是细胞分裂素信号转导通路的主要组成部分。
第十五章细胞信号转导 复习测试 （一）名词解释 1. 受体 2. 激素 3. 信号分子 4. G蛋白 5. 细胞因子 6. 自分泌信号传递 7. 蛋白激酶 8. 钙调蛋白 9. G蛋白偶联型受体 10. 向上调节 11. 细胞信号转导途径 12. 第二信使 （二）选择题 A型题： 1. 关于激素描述错误的是： A. 由内分泌腺/细胞合成并分泌 B. 经血液循环转运 C. 与相应的受体共价结合 D. 作用的强弱与其浓度相关 E. 可在靶细胞膜表面或细胞内发挥作用 2. 下列哪种激素属于多肽及蛋白质类： A. 糖皮质激素 B. 胰岛素 C. 肾上腺素 D. 前列腺素 E. 甲状腺激素 3. 生长因子的特点不包括： A. 是一类信号分子 B. 由特殊分化的内分泌腺所分泌 C. 作用于特定的靶细胞 D. 主要以旁分泌和自分泌方式发挥作用 E. 其化学本质为蛋白质或多肽 4. 根据经典的定义，细胞因子与激素的主要区别是： A. 是一类信号分子 B. 作用于特定的靶细胞 C. 由普通细胞合成并分泌 D. 可调节靶细胞的生长、分化 E. 以内分泌、旁分泌和自分泌方式发挥作用 5. 神经递质、激素、生长因子和细胞因子可通过下列哪一条共同途径传递信号：
A. 形成动作电位 B. 使离子通道开放 C. 与受体结合 D. 通过胞饮进入细胞 E. 自由进出细胞 6. 受体的化学本质是： A. 多糖 B. 长链不饱和脂肪酸 C. 生物碱 D. 蛋白质 E. 类固醇 7. 受体的特异性取决于： A. 活性中心的构象 B. 配体结合域的构象 C. 细胞膜的流动性 D. 信号转导功能域的构象 E. G蛋白的构象 8. 关于受体的作用特点，下列哪项是错误的： A. 特异性较高 B. 是可逆的 C. 其解离常数越大，产生的生物效应越大 D. 是可饱和的 E. 结合后受体可发生变构 9. 下列哪项与受体的性质不符： A. 各类激素有其特异性的受体 B. 各类生长因子有其特异性的受体 C. 神经递质有其特异性的受体 D. 受体的本质是蛋白质 E. 受体只存在于细胞膜上 10. 下列哪种受体是催化型受体： A. 胰岛素受体 B. 甲状腺激素受体 C. 糖皮质激素受体 受体 D. 肾上腺素能受体 E. 活性维生素D 3 11. 酪氨酸蛋白激酶的作用是： A. 使蛋白质结合上酪氨酸 B. 使含有酪氨酸的蛋白质激活 C. 使蛋白质中的酪氨酸激活 D. 使效应蛋白中的酪氨酸残基磷酸化 E. 使蛋白质中的酪氨酸分解 12. 下列哪种激素的受体属于胞内转录因子型： A. 肾上腺素 B. 甲状腺激素 C. 胰岛素 D. 促甲状腺素 E. 胰高血糖素
第十五章细胞信号转导 复习测试（一）名词解释 1. 受体 2. 激素 3. 信号分子 4. G蛋白 5. 细胞因子 6. 自分泌信号传递 7. 蛋白激酶 8. 钙调蛋白 9. G蛋白偶联型受体 10. 向上调节 11. 细胞信号转导途径 12. 第二信使 （二）选择题 A型题： 1. 关于激素描述错误的是： A. 由内分泌腺/细胞合成并分泌 B. 经血液循环转运 C. 与相应的受体共价结合 D. 作用的强弱与其浓度相关 E. 可在靶细胞膜表面或细胞内发挥作用 2. 下列哪种激素属于多肽及蛋白质类： A. 糖皮质激素 B. 胰岛素 C. 肾上腺素 D. 前列腺素 E. 甲状腺激素 3. 生长因子的特点不包括： A. 是一类信号分子 B. 由特殊分化的内分泌腺所分泌 C. 作用于特定的靶细胞 D. 主要以旁分泌和自分泌方式发挥作用
E. 其化学本质为蛋白质或多肽 4. 根据经典的定义，细胞因子与激素的主要区别是： A. 是一类信号分子 B. 作用于特定的靶细胞 C. 由普通细胞合成并分泌 D. 可调节靶细胞的生长、分化 E. 以内分泌、旁分泌和自分泌方式发挥作用 5. 神经递质、激素、生长因子和细胞因子可通过下列哪一条共同途径传递信号： A. 形成动作电位 B. 使离子通道开放 C. 与受体结合 D. 通过胞饮进入细胞 E. 自由进出细胞 6. 受体的化学本质是： A. 多糖 B. 长链不饱和脂肪酸 C. 生物碱 D. 蛋白质 E. 类固醇 7. 受体的特异性取决于： A. 活性中心的构象 B. 配体结合域的构象 C. 细胞膜的流动性 D. 信号转导功能域的构象 E. G蛋白的构象 8. 关于受体的作用特点，下列哪项是错误的： A. 特异性较高 B. 是可逆的 C. 其解离常数越大，产生的生物效应越大 D. 是可饱和的 E. 结合后受体可发生变构 9. 下列哪项与受体的性质不符： A. 各类激素有其特异性的受体 B. 各类生长因子有其特异性的受体 C. 神经递质有其特异性的受体 D. 受体的本质是蛋白质
细胞信号转导途径研究方法 一、蛋白质表达水平和细胞内定位研究 1、信号蛋白分子表达水平及分子量检测: Western blot analysis. 蛋白质印迹法是将蛋白质混合样品经SDS-PAGE后,分离为不同条带，其中含有能与特异性抗体(或McAb)相应的待检测的蛋白质(抗原蛋白)，将PAGE胶上的蛋白条带转移到NC 膜上此过程称为blotting，以利于随后的检测能够的进行，随后,将NC膜与抗血清一起孵育，使第一抗体与待检的抗原决定簇结合(特异大蛋白条带)，再与酶标的第二抗体反应,即检测样品的待测抗原并可对其定量。 基本流程： 检测示意图： 2、免疫荧光技术Immunofluorescence （IF） 免疫荧光技术是根据抗原抗体反应的原理，先将已知的抗原或抗体标记上荧光素制成荧光标记物，再用这种荧光抗体（或抗原）作为分子探针检查细胞或组织内的相应抗原（或抗体）。在细胞或组织中形成的抗原抗体复合物上含有荧光素，利用荧光显微镜观察标本，
荧光素受激发光的照射而发出明亮的荧光（黄绿色或桔红色），可以看见荧光所在的细胞或组织，从而确定抗原或抗体的性质、定位，以及利用定量技术测定含量。 采用流式细胞免疫荧光技术（FCM）可从单细胞水平检测不同细胞亚群中的蛋白质分子，用两种不同的荧光素分别标记抗不同蛋白质分子的抗体，可在同一细胞内同时检测两种不同的分子（Double IF），也可用多参数流式细胞术对胞内多种分子进行检测。 二、蛋白质与蛋白质相互作用的研究技术 1、免疫共沉淀（Co- Immunoprecipitation, Co-IP） Co-IP是利用抗原蛋白质和抗体的特异性结合以及细菌蛋白质的“protein A”能特异性地结合到免疫球蛋白的FC片段的现象而开发出来的方法。目前多用精制的protein A预先结合固化在agarose的beads上，使之与含有抗原的溶液及抗体反应后，beads上的prorein A 就能吸附抗原抗体达到沉淀抗原的目的。 当细胞在非变性条件下被裂解时，完整细胞内存在的许多蛋白质－蛋白质间的相互作用被保留了下来。如果用蛋白质X的抗体免疫沉淀X，那么与X在体内结合的蛋白质Y也能沉淀下来。进一步进行Western Blot和质谱分析。这种方法常用于测定两种目标蛋白质是否在体内结合，也可用于确定一种特定蛋白质的新的作用搭档。缺点：可能检测不到低亲和力和瞬间的蛋白质－蛋白质相互作用。 2、G ST pull-down assay GST pull-down assay是将谷胱甘肽巯基转移酶（GST）融合蛋白（标记蛋白或者饵蛋白，GST, His6, Flag, biotin …）作为探针，与溶液中的特异性搭档蛋白(test protein或者prey被扑获蛋白)结合，然后根据谷胱甘肽琼脂糖球珠能够沉淀GST融合蛋白的能力来确定相互作用的蛋白。一般在发现抗体干扰蛋白质－蛋白质之间的相互作用时，可以启用GST沉降技术。该方法只是用于确定体外的相互作用。 示意图：
第十一章细胞的信号转导 一、名词解释 1、细胞通讯 2、受体 3、第一信使 4、第二信使 5、G 蛋白 6、蛋白激酶A 二、填空题 1、细胞膜表面受体主要有三类即、、和。 2、在细胞的信号转导中，第二信使主要有、、、和。 3、硝酸甘油之所以能治疗心绞痛是因为它在体内能转化为，引起血管，从而减轻的负荷和的需氧量。 三、选择题 1、能与胞外信号特异识别和结合，介导胞内信使生成，引起细胞产生效应的是( )。 A、载体蛋白 B、通道蛋白 C、受体 D、配体 2、下列不属于第二信使的是（）。 A、cAMP B、cGMP C、DG D、CO 3、下列关于信号分子的描述中，不正确的一项是（）。 A、本身不参与催化反应 B、本身不具有酶的活性 C、能够传递信息 D、可作为酶作用的底物 4、生长因子是细胞内的（）。 A、结构物质 B、能源物质 C、信息分子 D、酶 5、肾上腺素可诱导一些酶将储藏在肝细胞和肌细胞中的糖原水解，第一个被激活的酶是（）。 A、蛋白激酶A B、糖原合成酶 C、糖原磷酸化酶 D、腺苷酸环化酶 6、（）不是细胞表面受体。 A、离子通道 B、酶连受体 C、G蛋白偶联受体 D、核受体 7、动物细胞中cAMP的主要生物学功能是活化（）。 A、蛋白激酶C B、蛋白激酶A C、蛋白激酶K D、Ca2+激酶 8、在G蛋白中，α亚基的活性状态是（）。 A、与GTP结合，与βγ分离 B、与GTP结合，与βγ聚合 C、与GDP结合，与βγ分离 D、与GDP结合，与βγ聚合
9、下面关于受体酪氨酸激酶的说法哪一个是错误的 A、是一种生长因子类受体 B、受体蛋白只有一次跨膜 C、与配体结合后两个受体相互靠近，相互激活 D、具有SH2结构域 10、在与配体结合后直接行使酶功能的受体是 A、生长因子受体 B、配体闸门离子通道 C、G蛋白偶联受体 D、细胞核受体 11、硝酸甘油治疗心脏病的原理在于 A、激活腺苷酸环化酶，生成cAMP B、激活细胞膜上的GC，生成cGMP C、分解生成NO，生成cGMP D、激活PLC，生成DAG 12、霍乱杆菌引起急性腹泻是由于 A、G蛋白持续激活 B、G蛋白不能被激活 C、受体封闭 D、蛋白激酶PKC功能异常 13下面由cAMP激活的酶是 A、PTK B、PKA C、PKC D、PKG 14下列物质是第二信使的是 A、G蛋白 B、NO C、GTP D、PKC 15下面关于钙调蛋白（CaM）的说法错误的是 A、是Ca2+信号系统中起重要作用 B、必须与Ca2+结合才能发挥作用 C、能使蛋白磷酸化 D、CaM激酶是它的靶酶之一16间接激活或抑制细胞膜表面结合的酶或离子通道的受体是 A、生长因子受体 B、配体闸门离子通道 C、G蛋白偶联受体 D、细胞核受体 17重症肌无力是由于 A、G蛋白功能下降
生物膜与细胞信号转导 名词解释： 1.脂筏：胆固醇分子不可能在脂双层里均匀分布，而是与鞘脂一起集中在膜的 特定区域，胆固醇-鞘脂漂浮在液态磷酸甘油脂“海洋”上的“筏”一样称为脂筏。 2.转运蛋白： 3.P-型ATPase：是阳离子转运蛋白，在转运过程中需要ATP可逆磷酸化的过 程，磷酸化使得转运蛋白的构象发生变化，同时，转运阳离子做跨膜运输4.次级主动运输：第一种溶质(S1)通过初级主动运输产生浓度梯度后，接着， 第一种溶质顺着浓度梯度提供能量，驱动第二种溶质(S2)逆浓度梯度运输 5.G蛋白分子开关：GTP酶(GTPase)是一个分子开关，开关是通过结合和水解 GTP进行控制。 6.激酶锚定蛋白：AKAPs 是支架蛋白，位于脂筏的胞质侧，将信号通路中执 行功能的蛋白聚集在一起，便于反应进行 7.信号蛋白 8.MAP激酶级联反应：酵母中的mating pheromones，果蝇中复眼的光受体的 分化，开花植物中对病源的防御反应。 简答题： 1.溶质分子跨膜运动，有哪几种机制？ 2.以细菌KcsA钾离子通道为例，说明电压门控的钾离子通道结构与运输的关系答：细菌KcsA通道是由四个亚基组成，其中两个亚基由两个跨膜的螺旋(M1和M2) 和通道胞外的孔区域(P)组成。每个亚基的M2螺旋线与另一个亚基的M2相互交叉形成一个“螺旋束”，封闭了面向质膜的孔，则K+不能通过；M2 螺旋线可以在具有甘氨酸残基的位点弯曲，将通道门打开。K+运输。 P区域是由一个长约1/3通道宽度的短的螺旋和一个能形成“衬里的”狭窄的选择性过滤器的无螺旋的环，允许K+通过。选择性过滤器的衬里含有高度保守的五肽骨架，产生5个连续排列的氧原子环。每个环由四个氧原子组成，直径是3nm，而失水K+直径是2.7nm。当通道门打开，K+进入通道时，电负性的氧原子替代了与K+结合的水分子，与K+稳定的相互作用使K+运输。尽管选择性过滤器具有4个K+结合位点，但实际上只能同时结合2个K+。 3.乙酰胆碱受体门控通道结构及离子运输机制 结构：乙酰胆碱是由运动神经元释放到肌细胞质膜，与乙酰胆碱受体结合，它可以改变受体的构象，引起离子通道打开。乙酰胆碱受体允许Na+、Ca2+和K+通过。由5个亚基组成：γ，β和δ各1个，2个α亚基，每个α亚基带有1个乙酰胆碱结合位点。每个亚基含有4个跨膜双螺旋，5个亚基围成1个中心孔，孔直径约20 ?，突出在胞质和细胞表面。 机制：2个乙酰胆碱结合到2个α亚基上，引起构象发生变化，使疏水侧链远离通道的中心，打开离子通道，让离子通过。组成5个亚基的M2螺旋所含有的5个Leu 侧链突出在通道，限制了通道的直径。当两个乙酰胆碱受体位点被占据，构想发生变化，随着M2螺旋的轻微扭曲，5个Leu残基旋转，远离通道中心，由较小的极性氨基酸代替，通过道门打开，允许Ca2+，Na+、K+通过。
细胞受体类型、特点 及重要的细胞信号转导途径 学院：动物科学技术学院 专业：动物遗传育种与繁殖 姓名：李波
学号：2015050509
目录 1、细胞受体类型及特点 (4) 1.1离子通道型受体 (4) 1.2 G蛋白耦联型受体 (4) 1.3 酶耦联型受体 (5) 2、重要的细胞信号转导途径 (5) 2.1细胞内受体介导的信号传递 (5) 2.2 G蛋白偶联受体介导的信号转导 (6) 2.2.1激活离子通道的G蛋白偶联受体所介导的信号通路 (7) 2.2.2激活或抑制腺苷酸环化酶的G蛋白偶联受体 (7) 2.2.3 激活磷脂酶C、以lP3和DAG作为双信使 G蛋白偶联受体介导的信号通 路 (8) 2.2 酶联受体介导的信号转导 (9) 2.2.1 受体酪氨酸激酶及RTK-Ras蛋白信号通路 (10) 2.2.2 P13K-PKB(Akt)信号通路 (10) 2.2.3 TGF-p—Smad信号通 (11) 2.2.4 JAK—STAT信号通路 (12)
1、细胞受体类型及特点 受体(receptor)是一种能够识别和选择性结合某种配体(信号分子)的大分子物质，多为糖蛋白，一般至少包括两个功能区域，与配体结合的区域和产生效应的区域，当受体与配体结合后，构象改变而产生活性，启动一系列过程，最终表现为生物学效应。受体与配体问的作用具有3个主要特征：①特异性；②饱和性；③高度的亲和力。 根据靶细胞上受体存在的部位，可将受体分为细胞内受体(intracellular receptor)和细胞表面受体(cell surface receptor)。细胞内受体介导亲脂性信号分子的信息传递，如胞内的甾体类激素受体。细胞表面受体介导亲水性信号分子的信息传递，膜表面受体主要有三类：①离子通道型受体(ion—channel—linked receptor)；②G蛋白耦联型受体(G—protein —linked receptor)；③酶耦联的受体(enzyme—linked recep—tor)。第一类存在于可兴奋细胞。后两类存在于大多数细胞，在信号转导的早期表现为激酶级联事件，即为一系列蛋白质的逐级磷酸化，借此使信号逐级传送和放大。 1.1离子通道型受体 离子通道型受体是一类自身为离子通道的受体，即配体门通道(1igand—gated channel)，主要存在于神经、肌肉等可兴奋细胞，其信号分子为神经递质。神经递质通过与受体的结合而改变通道蛋白的构象，导致离子通道的开启或关闭，改变质膜的离子通透性，在瞬间将胞外化学信号转换为电信号，继而改变突触后细胞的兴奋性。如：乙酰胆碱受体以三种构象存在，两分子乙酰胆碱的结合可以使之处于通道开放构象，但该受体处于通道开放构象状态的时限仍十分短暂，在几十毫微秒内又回到关闭状态。然后乙酰胆碱与之解离，受体则恢复到初始状态，做好重新接受配体的准备。离子通道型受体分为阳离子通道，如乙酰胆碱、谷氨酸和五羟色胺的受体，和阴离子通道。 1.2 G蛋白耦联型受体 三聚体GTP结合调节蛋白(trimeric GTP—binding regulatory protein)简称G蛋白，位于质膜胞质侧，由a、p、-／三个亚基组成，a和7亚基通过共价结合的脂肪酸链尾结合在膜上，G蛋白在信号转导过程中起着分子开关的作用，当a亚基与GDP结合时处于关闭状态，与GTP结合时处于开启状态，“亚基具有GTP酶活性，能催化所结合的ATP 水解，恢复无活性的三聚体状态，其GTP酶的活性能被RGS(regulator of G protein signaling)增强。RGS也属于GAP(GTPase activating protein)。 G蛋白耦联型受体为7次跨膜蛋白(图10—6)，受体胞外结构域识别胞外信号分子并与之结合，胞内结构域与G蛋白耦联。通过与G蛋白耦联，调节相关酶活性，在细胞内
表皮生长因子两条典型的信号转导途径 (1)表皮生长因子受体介导的信号转导途径 表皮生长因子与其受体－表皮生长因子受体结合后可引发一系列细胞内变化，最终使细胞发生分化或增殖。表皮生长因子受体是一种受体酪氨酸蛋白激酶，而受体酪氨酸蛋白激酶→Ras→MAPK级联途径是表皮生长因子刺激信号传递到细胞核内的最主要途径。它由以下成员组成：表皮生长因子受体→含有SH2结构域的接头蛋白(如Grb2)→鸟嘌呤核苷酸释放 因子(如SOS)→Ras蛋白→MAPKKK(如Raf1)→MAPKK→MAPK→转录因子等(图21－24)。 表皮生长因子与受体结合后，可以使受体发生二聚体化，从而改变了受体的构象，使其中的蛋白酪氨酸激酶活性增强，受体自身的酪氨酸残基发生磷酸化，磷酸化的受体便形成了与含SH2结构域的蛋白分子Grb2结合的位点，导致Grb2与受体的结合。Grb2中有两个SH3结构域，该部位与一种称为SOS的鸟苷酸交换因子结合，使之活性改变，SOS则进一步活化Ras，激活的Ras作用于MAPK激活系统，导致ERK的激活。最后ERK转移到 细胞核内，导致某些转录因子的活性改变从而改变基因的表达状态及细胞的增殖与分化过程。 (2)γ－干扰素受体介导的信号转导 γ－干扰素是由活化T细胞产生的，它具有促进抗原提呈和特异性免疫识别的作用，并可促进B细胞分泌抗体。γ－干扰素与受体结合以后，也可以导致受体二聚体化，二聚体化的受体可以激活JAK－STAT系统，后者将干扰素刺激信号传入核内。JAK(Janus Kinase)为一种存在于胞浆中的蛋白酪氨酸激酶，它活化后可使干扰素受体磷酸化。STAT(Signal Transducerand Activator of Transcription)可以通过其SH2结构域识别磷酸化的受体并与之结合。然后STAT分子亦发生酪氨酸的磷酸化，酪氨酸磷酸化的STAT进入胞核形成有活性的转录因子，影响基因的表达 表皮生长因子（epidermal growth factor，EGF）受体是研究得比较清楚的酪氨酸激酶受体，存在于特殊的靶细胞的质膜上，调节不同的功能，包括细胞的生长、增殖和分化，并且与肿瘤的发生有关。EGF受体（EGF receptor）结构是一种糖蛋白，由三个部分组成：①细胞外结构域有621个氨基酸残基，富含半胱氨酸，并形成多对二硫键。其上结合有糖基，是EGF结合的位点；②跨膜区由23个氨基酸残基组成；③细胞质结构域，由542个氨基酸残基组成，含有无活性的酪氨酸激酶和几个酪氨酸磷酸化的位点。当EGF同受体细胞外结构域结合位点结合后，受体被激活，导致两个EGF受体单体形成二聚体，激活细胞质部分的酪氨酸激酶，使酪氨酸自我磷酸化。EGF受体上有五个主要的磷酸化的酪氨酸位点，可以同几种不同的蛋白质结合，分别引起细胞内不同的信号应答。在多数情况下，EGF受体被磷酸化的酪氨酸位点同靶蛋白（酶）的SH2结构域相互作用，将靶蛋白（酶）激活，引起细胞应答。如PIP2激酶通过SH2与EGF受体磷酸化的酪氨酸位点相互作用被激活，激活的PIP2激酶使一种膜脂-PIP2磷酸化。另外，磷酸化的酪氨酸位点也可以同具有SH2结构域的磷脂酶Cγ相互作用，并将磷脂酶Cγ激活，激活的磷脂酶Cγ可将质膜中PIP2水解成IP3和DAG，引起与磷脂肌醇-G蛋白偶联系统类似的信号转导。
第三节主要的信号转导途径 一、膜受体介导的信号传导 （一）cAMP-蛋白激酶A途径 述：该途径以靶细胞内cAMP浓度改变和激活蛋白激酶A（PKA）为主要特征，是激素调节物质代谢的主要途径。 1．cAMP的合成与分解 ⑴引起cAMP水平增高的胞外信号分子：胰高血糖素、肾上腺素、 促肾上腺皮质激素、促甲状腺素、甲状旁腺素和加压素等。 α-GDP-βγ（Gs蛋白）激素+受体→激素-受体→↓ α-GTP + βγ ↓ AC激活 ↓ ATP →cAMP 述：当信号分子（胰高血糖素、肾上腺素和促肾上腺皮质激素）与靶细胞质膜上的特异性受体结合，形成激素一受体复合物 而激活受体。活化的受体可催化Gs的GDP与GTP交换，导 致Gs的α亚基与βγ解离，蛋白释放出αs-GTP。αs-GTP能激 活腺苷酸环化酶，催化ATP转化成cAMP，使细胞内cAMP 浓度增高。过去认为G蛋白中只有α亚基发挥作用，现知βγ 复合体也可独立地作用于相应的效应物，与α亚基拮抗。 腺苷酸环化酶分布广泛，除成熟红细胞外，几乎存在于所有组织的细胞质膜上。cAMP经磷酸二酯酶（PDE）降解成 5'－AMP而失活。cAMP是分布广泛而重要的第二信使。
⑵AC活性的抑制与cAMP浓度降低 ◇Gα-GTP结合AC并使之激活后，同时激活自身的GTP酶活性，Gα-GTP→Gα-GDP，Gs、AC均失活。从而在细胞对cAMP浓度升高作出应答后AC活性迅速逆转。 ⑶少数激素，如生长激素抑制素、胰岛素和抗血管紧张素II 等，它们活化受体后可催化抑制性G蛋白解离，导致细胞内AC活性下降，从而降低细胞内cAMP水平。 ⑷正常细胞内cAMP的平均浓度为10－6mol/L。cAMP在细 胞中的浓度除与腺苷酸环化酶活性有关外，还与磷酸二酯酶的活性有关。举例如下： ①一些激素如胰岛素，能激活磷酸二酯酶，加速cAMP降解； ②某些药物如茶碱，则抑制磷酸二酯酶，促使细胞内cAMP 浓度升高。 2．cAMP的作用机制――cAMP激活PKA（幻灯64） ⑴cAMP对细胞的调节作用是通过激活cAMP依赖性蛋白激酶 或称蛋白激酶A （PKA）系统来实现的。 ⑵PKA的结构 2C（催化亚基）：蛋白丝/苏氨酸磷酸化酶活性四聚体蛋白 变构酶 2R（调节亚基）：各有2个cAMP结合位点述：催化亚基有催化底物蛋白质某些特定丝／苏氨酸残基磷酸化的功能。调节亚基与催化亚基相结合时，PKA呈无活性状态。当4分子cAMP与2个调节亚基结合后，调节亚基脱落，游离的催化亚基具有蛋白激酶活性。PKA的激活过程需要Mg2+。
第八章细胞信号转导 名词解释 1、蛋白激酶protein kinase 将磷酸基团转移到其他蛋白质上的酶，通常对其他蛋白质的活性具有调节作用。 2、蛋白激酶C protein kinase C 一类多功能的丝氨酸/苏氨酸蛋白激酶家族，可磷酸化多种不同的蛋白质底物。 3、第二信使second messenger 第一信使分子（激素或其他配体）与细胞表面受体结合后，在细胞内产生或释放到细胞内的小分子物质，如cAMP，IP3，钙离子等，有助于信号向胞内进行传递。 4、分子开关molecular switch 细胞信号转导过程中，通过结合GTP与水解GTP，或者通过蛋白质磷酸化与去磷酸化而开启或关闭蛋白质的活性。 5、磷脂酶C phospholipid C 催化PIP2分解产生1,4,5-肌醇三磷酸（IP3）和二酰甘油(DAG)两个第二信使分子。 6、门控通道gated channel 一种离子通道，通过构象改变使溶液中的离子通过或阻止通过。依据引发构象改变的机制的不同，门控通道包括电位门通道和配体门通道两类。 7、神经递质neurotransmitter 突触前端释放的一种化学物质，与突触后靶细胞结合，并改变靶细胞的膜电位。 8、神经生长因子nerves growth factor，NGF 神经元存活所必需的细胞因子 9、受体receptor 任何能与特定信号分子结合的膜蛋白分子，通常导致细胞摄取反应或细胞信号转导。10、受体介导的胞吞作用receptor mediated endocytosis 通过网格蛋白有被小泡从胞外基质摄取特定大分子的途径。被转运的大分子物质与细胞表面互补性的受体结合，形成受体-配体复合物并引发细胞质膜局部内化作用，然后小窝脱离质膜形成有被小泡而将物质吞入细胞内。 11、受体酪氨酸激酶receptor tyrosine kinase，RTK 能将自身或胞质中底物上的酪氨酸残基磷酸化的细胞表面受体。主要参与细胞生长和分化的调控。 12、调节型分泌regulated secretion 细胞中已合成的分泌物质先储存在细胞质周边的分泌泡中，在受到适宜的信号刺激后，才与质膜融合将内容物分泌到细胞表面。 13、细胞通讯cell communication 信号细胞发出的信息传递到靶细胞并与受体相互作用，引起靶细胞产生特异性生物学效应的过程。 14、细胞信号传递cell signaling 通过信号分子与受体的相互作用，将外界信号经细胞质膜传递到细胞内部，通常传递至细胞核，并引发特异性生物学效应的过程。 15、信号转导signal transduction 细胞将外部信号转变为自身应答反应的过程。 16、组成型分泌constitutivesecretion
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PPAR-γ作用及其相关信号转导途径 作者：陈永熙， 王伟铭， 周同， 陈楠， Yong-Xi Chen， Wei-Ming Wang， Tong Zhou， Nan Chen 作者单位：上海交通大学医学院附属瑞金医院肾内科,上海,200032 刊名： 细胞生物学杂志 英文刊名：CHINESE JOURNAL OF CELL BIOLOGY 年，卷(期)：2006,28(3) 被引用次数：86次 参考文献(43条) 1.Rocchi S查看详情 1999 2.Sundvold H查看详情 2001 3.Rosen ED查看详情 2002 4.Fajas L查看详情 1998 5.Tautenhahn A查看详情 2003 6.Werman A查看详情 1997 7.Hah J查看详情 2000 8.Hsi LC查看详情 2001 9.Nolte RT查看详情 1998 10.Zingarelli B查看详情 2005 11.Li M查看详情 2000 12.Li Q查看详情 2002(02) 13.Bohrer H查看详情 1997 14.Ghosh S查看详情 1998 15.Straus DS查看详情 2000 16.Chung SW查看详情 2000 17.Sanchez-Hidalgo M查看详情 2005 18.Desvergne B查看详情 1999 19.Guan Y查看详情 2001 20.Tontonoz P查看详情 1998 21.Bruemmer D查看详情 2003 22.Delerive P查看详情 1999 23.Marx N查看详情 1999 24.Verrier E查看详情 2004 25.Fan WH查看详情 2000 26.Fu M查看详情 2001 27.Bendixen AC查看详情 2001 28.Goetze S查看详情 2002 29.Anandharajan R查看详情 2005 30.Hegyi K查看详情 2004
第十九章细胞信息转导的分子机制一、A型选择题 1．不作用于质膜受体的信息物质是 A．乙酰胆碱 D．甲状腺素 2．能激活PKG的是 A．cAMP A．脂类质 4．下列哪种物质能使蛋白质的酪氨酸残基发生磷酸化A．PKA B．PKC C．生长激素受体 D．类固醇激素受体 A．1个 A．1个 E．糖皮质激素受体 C．3个 C．3个 D．4个 D．4个 E．5个 E．5个 E．失活 B．cGMP B．糖类 C．Ca2+ D．DAG D．多肽 E．GTP E．蛋白 3．细胞膜受体的本质是 C．核酸 B．谷氨酸 E．神经酰胺 C．表皮生长因子 5．PKA中的每个调节亚基可结合cAMP的分子数为B．2个 B．2个
6．PKA所含的亚基数为 7．蛋白激酶的作用是使蛋白质或酶 A．磷酸化 B．脱磷酸化 C．水解 8．通过膜受体起调节作用的激素是 A．性激素 B．糖皮质激素 C．甲状腺素 9．下列关于GTP结合蛋白(G蛋白)的叙述,错误的是 （2007年全国硕士研究生入学考试西医综合科目试题） A．膜受体通过G蛋白与腺苷酸环化酶偶联 B．可催化GTP水解为GDP C．霍乱毒素可使其失活 D．有三种亚基α、β、γ D．激活 D．肾上腺素 E．活性维生素 E．G蛋白具有内源GTP酶活性 10．下列哪种酶激活后会直接引起cAMP浓度降低 （2006年全国硕士研究生入学考试西医综合科目试题） A．蛋白激酶A B．蛋白激酶C D．磷脂酶C E．蛋白激酶G 11．cAMP能别构激活下列哪种酶（2005年全国硕士研究生入学考试西医综合科目试题） A．磷脂酶A B．蛋白激酶A D．蛋白激酶G E．酪氨酸蛋白激酶 12．细胞膜内外正常Na+和K+浓度差的形成和维持是由于 （2004年全国硕士研究生入学考试西医综合科目试题） A．膜安静时K+通透性大 C．Na+易化扩散的结果 C．磷酸二酯酶 C．蛋白激酶C B．膜兴奋时Na+通透性增加 D．膜上Na+泵的作用 E．膜上Ca2+泵的作用