植物中钙依赖蛋白激酶（CDPKs）的结构与功能

在植物细胞中,钙离子作为第二信使,通过钙依赖蛋白激酶（CDPKs)发挥功能是其传递信号的主要途径之一。CDPKs广泛存在于植物体中,是目前植物体内研究最深入的蛋白激酶,在简要阐述CDPKs于植物体内的分布定位的基础上,介绍了CDPKs的结构特点,生化性质及其在植物细胞生理功能中的作用,并就该领域的研究前景作了展望。     

钙依赖的蛋白激酶与植物抗逆性

植物钙依赖的蛋白激酶(Calcium-dependent protein kinases,CDPKs)是细胞Ca2 信号的受体,同时具有Ca2 受体蛋白和Ser/Thr蛋白激酶的功能。许多植物CDPKs基因受环境胁迫刺激发生表达水平的改变,这些基因在植物逆境胁迫的Ca2 信号转导中起着十分重要的作用,为植物CDPKs抗逆功能的研究和植物的抗逆遗传改良提供了理论基础和基因资源。     

钙依赖蛋白激酶（CDPKs）在植物钙信号转导中的作用

CDPKs在植物钙信号转导中起重要作用。本文介绍了植物钙信号转导及CDPKs的结构与生化性质,在此基础上,重点总结了CDPKs在植物钙信号转导中的潜在调节作用,包括基因表达、代谢、离子和水分的跨膜运输、细胞骨架的动态变化、气孔运动和生长发育等,并提出了在CDPKs研究中已达成的共识和需要解决的问题。     

植物钙感受器及其介导的逆境信号途径

钙信号是植物生长发育和逆境响应的重要调控因子, 是植物生理与逆境生物学研究领域中的热点之一。当植物细胞受到外界逆境刺激时, 其胞内会产生具有时空特异性的Ca2+信号变化, 这种变化首先被胞内钙感受器所感知并解码, 再由钙感受器互作蛋白将信号传递到下游, 从而激活下游早期响应基因的表达或相关离子通道的活性, 最终产生特异性逆境响应。植物细胞通过感知胞内钙信号的变化如何识别来自外界不同性质或不同强度的刺激, 是近几年植物生物学家所关注的科学问题。文章主要总结了近几年在植物钙感受器研究领域中的最新进展, 包括钙依赖蛋白激酶(CDPKs)、钙调素(CaMs)、类钙调素蛋白(CMLs)、类钙调磷酸酶B蛋白(CBLs)及其互作蛋白激酶(CIPKs)等的结构、功能及其介导的逆境信号途径, 并提供新的见解和展望。     

钙依赖蛋白激酶(CDPKs)在植物钙信号转导中的作用

CDPKs在植物钙信号转导中起重要作用。本文介绍了植物钙信号转导及CDPKs的结构与生化性质,在此基础上,重点总结了CDPKs在植物钙信号转导中的潜在调节作用,包括基因表达、代谢、离子和水分的跨膜运输、细胞骨架的动态变化、气孔运动和生长发育等,并提出了在CDPKs研究中已达成的共识和需要解决的问题。     

三种隐孢子虫钙依赖蛋白激酶的生物信息学分析

对三种隐孢子虫(C.parvum Iowa II、C.hominis TU502和C.muris RN66)钙依赖蛋白激酶(Calcium-dependent protein kinases,CDPKs)进行生物信息学分析,探索该蛋白的结构并预测其功能,为其基因功能的研究提供一定的理论基础。通过隐孢子虫基因组数据库收集数据,获得三种隐孢子虫CDPKs蛋白的序列信息,通过生物信息学软件进行分析,预测该蛋白的理化性质、翻译后修饰位点、功能域、亚细胞定位、二级结构、亲/疏水性、抗原表位等。隐孢子虫CDPKs的蛋白性质不稳定,理论分子量从59.76 k Da到76.63 k Da,p I值为5.33~6.09,CDPKs不具有跨膜区和信号肽,不是跨膜分泌性蛋白,都具有蛋白激酶C磷酸化位点、酪氨酸激酶Ⅱ磷酸化位点、酪氨酸激酶磷酸化位点、c AMP和c GMP依赖蛋白激酶磷酸化位点、N-端糖基化位点、N-端肉豆蔻酰化位点和EF-hand钙结合域,二级结构主要以α螺旋和无规卷曲为主;CDPKs主要存在虫体细胞内,均有20多个潜在的抗原表位。在隐孢子虫中,CDPKs蛋白不仅可单独发挥作用,而且还能通过相互结合发挥其生物学效应;同时,CDPKs有望成为候选疫苗及潜在药物靶点。     

大花杓兰钙依赖蛋白激酶基因克隆及植物表达载体构建

RT-PCR结合RACE技术,克隆到1个全长2079 bp的大花杓兰钙依赖蛋白激酶基因CmCDPK,cDNA为1491 bp,编码496个氨基酸。CmCDPK是1个具有CDPKs典型的Ser/Thr蛋白激酶保守结构域、含1个跨膜结构域、无信号肽、稳定的亲水性蛋白。CmCDPK二级结构主要由α-螺旋和无规卷曲构成。相对于其他植物CDPKs,CmCDPK与小兰屿蝴蝶兰和铁皮石斛的亲缘关系更近。通过DNA重组技术将CmCDPK片段克隆到pBI121质粒上。PCR、酶切及DNA测序的结果表明,重组质粒pBI-CmCDPK包含1个1491 bp的CmCDPK片段,且核苷酸序列及插入方向完全正确。本研究首次克隆了大花杓兰CmCDPK基因,并成功构建了植物过表达载体pBI-CmCDPK,为CmCDPK基因在烟草中实现遗传转化和功能研究奠定基础。     

利用酵母双杂交系统筛选水稻中与OsCRK5互作蛋白

水稻是重要的粮食作物,研究水稻生长发育调控机制可为水稻品种改良奠定理论基础。钙依赖蛋白激酶（calcium dependent protein kinases, CDPKs）是植物中重要的蛋白激酶,参与植物生长发育以及对环境反应的应答。水稻中的CRK5(CDPKrelated kinase 5)在蛋白序列和结构上与CDPK高度同源。为进一步研究OsCRK5在水稻干旱反应中的功能,本研究利用酵母双杂交筛库技术筛选了OsCRK5的互作蛋白。首先将OsCRK5的1-1 332 bp片段克隆至pGBKT7载体中,获得诱饵载体pGBKT7-OsCRK5,经测序无误后,转化至酵母菌株Y2H Gold中。在营养缺陷培养基中观察到重组蛋白不具有毒性作用及自激活活性,同时利用Western blot分析重组蛋白的表达。进一步利用水稻cDNA文库筛选OsCRK5的互作蛋白,共得到77个阳性克隆。功能预测结果显示,互作蛋白涉及蛋白质合成、贮存和降解过程、转录调控、植物细胞生长和分裂过程、能量代谢和细胞代谢等方面的功能。最后,从阳性克隆中选取参与植物干旱胁迫应答的两个蛋白OsWR1和OsDi19-1,利用酵母...     

植物体内蛋白质磷酸化及其在信息传递中的作用

本文概述了蛋白质磷酸化作用及其在细胞信息传递中的作用以及目前植物体内发现的蛋白激酶类以及经受磷酸化作用的蛋白质和酶类。     

植物蛋白激酶介导的非生物胁迫和激素信号转导途径的研究进展

干旱、盐渍、低温和高温等非生物胁迫严重影响植物的生长发育和作物的产量。在长期的进化过程中,植物逐渐形成了对外部刺激快速感知和主动适应的能力,其中植物体内逆境信号的传递在植物快速感知外部刺激和主动适应非生物胁迫过程中起着非常重要的作用。蛋白激酶和蛋白磷酸酶催化的蛋白质磷酸化和去磷酸化是植物体内存在的最普遍且最重要的信号转导调节方式。其中,蛋白激酶的主要作用是将ATP或GTP上的γ磷酸基团转移到特定的底物蛋白上,使蛋白磷酸化,被磷酸化的蛋白发挥相应的生理功能。近年来,利用生物技术和基因工程等手段从细胞、分子水平上研究有关蛋白激酶的抗逆机理,通过基因沉默、基因过表达等策略提高植物的抗逆性成为国内外抗逆分子生物学与分子育种学研究的热点。本文主要对植物蛋白激酶在介导非生物胁迫和激素信号通路中的作用进行综述,为进一步研究植物蛋白激酶功能提供有价值的信息。     

Molecular evolution of calmodulin-like domain protein kinases (CDPKs) in plants and protists

Many genes for calmodulin-like domain protein kinases (CDPKs) have been identified in plants and Alveolate protists. To study the molecular evolution of the CDPK gene family, we performed a phylogenetic analysis of CDPK genomic sequences. Analysis of introns supports the phylogenetic analysis; CDPK genes with similar intron/exon structure are grouped together on the phylogenetic tree. Conserved introns support a monophyletic origin for plant CDPKs, CDPK-related kinases, and phosphoenolpyruvate carboxylase kinases. Plant CDPKs divide into two major branches. Plant CDPK genes on one branch share common intron positions with protist CDPK genes. The introns shared between protist and plant CDPKs presumably originated before the divergence of plants from Alveolates. Additionally, the calmodulin-like domains of protist CDPKs have intron positions in common with animal and fungal calmodulin genes. These results, together with the presence of a highly conserved phase zero intron located precisely at the beginning of the calmodulin-like domain, suggest that the ancestral CDPK gene could have originated from the fusion of protein kinase and calmodulin genes facilitated by recombination of ancient introns. Received: 11 July 2000 / Accepted: 18 April 2001     

CDPK-mediated abiotic stress signaling

Calcium-dependent protein kinases (CDPKs) constitute a large multigene family in various plant species. CDPKs have been shown to have important roles in various physiological processes, including plant growth and development and abiotic and biotic stress responses in plants. Functional analysis using gain-of-function and loss-of-function mutants has revealed the biological function of CDPKs in planta. Several CDPKs have been shown to be essential factors in abiotic stress tolerance, positively or negatively regulating stress tolerance by modulating ABA signaling and reducing the accumulation of reactive oxygen species (ROS). This review summarizes recent results describing the biological function of CDPKs that are involved in abiotic stress tolerance.     

Calcium dependent protein kinase 1 and calcium fluxes in the malaria parasite

Calcium dependent protein kinases (CDPKs) are found only in plants and alveolates and are distinguished from other kinases by an activation domain that binds calcium directly. Plants contain families of these kinases and their functions are modulated by post translational modifications as well as calcium activation. Apicomplexan parasites also contain CDPK families and this review is focused on CDPK1 in Plasmodium spp. This enzyme has been implicated in parasite motility and host cell invasion and at least two substrates associated with the actomyosin motor complex have been identified. By analogy with the plant CDPKs we propose that its activity is modulated both by post translational modifications and by its subcellular location in a compartment within the parasite's pellicle, which may regulate the calcium concentration required for activation.     

Identification of a novel phosphorylation motif for CDPKs: phosphorylation of synthetic peptides lacking basic residues at P-3/P-4

The Ca(2+)-dependent protein kinases (CDPKs) are members of a large subfamily of protein kinases in plants that have been implicated in the control of numerous aspects of plant growth and development. One known substrate of the CDPKs is the ER-located ACA2 calcium pump, which is regulated by phosphorylation of Ser(45). In the present study, a synthetic peptide based on the known regulatory phosphorylation site (RRFRFTANLS(45)KRYEA) was efficiently phosphorylated in vitro by CDPKs but not a plant SNF1-related protein kinase. Phosphorylation of the Ser(45)-ACA2 peptide was surprising because the sequence lacks basic residues at P-3/P-4 (relative to the phosphorylated Ser at position P) that are considered to be essential recognition elements for CDPKs. We demonstrate that phosphorylation of the Ser(45)-ACA2 peptide is dependent on the cluster of basic residues found N-terminal (P-6 to P-9) as well as C-terminal (P + 1/P + 2) to the phosphorylated Ser. The results establish a new general phosphorylation motif for CDPKs: [Basic-Basic-X-Basic]-phi-X(4)-S/T-X-Basic (where phi is a hydrophobic residue). The motif predicts a number of new phosphorylation sites in plant proteins. Evidence is presented that the novel motif may explain the phosphorylation by CDPKs of Ser271 in the aquaporin PM28A.     

Calcium signaling through protein kinases. The Arabidopsis calcium-dependent protein kinase gene family

In plants, numerous Ca(2+)-stimulated protein kinase activities occur through calcium-dependent protein kinases (CDPKs). These novel calcium sensors are likely to be crucial mediators of responses to diverse endogenous and environmental cues. However, the precise biological function(s) of most CDPKs remains elusive. The Arabidopsis genome is predicted to encode 34 different CDPKs. In this Update, we analyze the Arabidopsis CDPK gene family and review the expression, regulation, and possible functions of plant CDPKs. By combining emerging cellular and genomic technologies with genetic and biochemical approaches, the characterization of Arabidopsis CDPKs provides a valuable opportunity to understand the plant calcium-signaling network.     

Protein kinases in plant responses to drought,salt, and cold stress

Protein kinases are major players in various signal transduction pathways. Understanding the molecular mechanisms behind plant responses to biotic and abiotic stresses has become critical for developing and breeding climate-resilient crops. In this review,we summarize recent progress on understanding plant drought, salt, and cold stress responses, with a focus on signal perception and transduction by different protein kinases, especially sucrose nonfermenting1(SNF1)-related protein kinases(Sn RKs),mitogen-activated protein kinase(MAPK) cascades,calcium-dependent protein kinases(CDPKs/CPKs),and receptor-like kinases(RLKs). We also discuss future challenges in these research fields.     

Unusual Membrane-Associated Protein Kinases in Higher Plants

Plant genomes encode a variety of protein kinases, and while some are functional homologues of animal and fungal kinases, others have a novel structure. This review focuses on three groups of unusual membrane-associated plant protein kinases: receptor-like protein kinases (RLKs), calcium-dependent protein kinases (CDPKs), and histidine protein kinases. Animal RLKs have a putative extracellular domain, a single transmembrane domain, and a protein kinase domain. In plants, all of the RLKs identified thus far have serine/threonine signature sequences, rather than the tyrosine-specific signature sequences common to animals. Recent genetic experiments reveal that some of these plant kinases function in development and pathogen resistance. The CDPKs of plants and protozoans are composed of a single polypeptide with a protein kinase domain fused to a C-terminal calmodulin-like domain containing four calcium-binding EF hands. No functional plant homologues of protein kinase C or Ca²⁺/calmodulin-dependent protein kinase have been identified, and no animal or fungal CDPK homologues have been identified. Recently, histidine kinases have been shown to participate in signaling pathways in plants and fungi. ETR1, an Arabidopsis histidine kinase homologue with three transmembrane domains, functions as a receptor for the plant hormone ethylene. G-protein-coupled receptors, which often serve as hormone receptors in animal systems, have not yet been identified in plants. Received: 18 August 1997/Revised: 23 December 1997     

The Arabidopsis CDPK-SnRK superfamily of protein kinases

The CDPK-SnRK superfamily consists of seven types of serine-threonine protein kinases: calcium-dependent protein kinase (CDPKs), CDPK-related kinases (CRKs), phosphoenolpyruvate carboxylase kinases (PPCKs), PEP carboxylase kinase-related kinases (PEPRKs), calmodulin-dependent protein kinases (CaMKs), calcium and calmodulin-dependent protein kinases (CCaMKs), and SnRKs. Within this superfamily, individual isoforms and subfamilies contain distinct regulatory domains, subcellular targeting information, and substrate specificities. Our analysis of the Arabidopsis genome identified 34 CDPKs, eight CRKs, two PPCKs, two PEPRKs, and 38 SnRKs. No definitive examples were found for a CCaMK similar to those previously identified in lily (Lilium longiflorum) and tobacco (Nicotiana tabacum) or for a CaMK similar to those in animals or yeast. CDPKs are present in plants and a specific subgroup of protists, but CRKs, PPCKs, PEPRKs, and two of the SnRK subgroups have been found only in plants. CDPKs and at least one SnRK have been implicated in decoding calcium signals in Arabidopsis. Analysis of intron placements supports the hypothesis that CDPKs, CRKs, PPCKs and PEPRKs have a common evolutionary origin; however there are no conserved intron positions between these kinases and the SnRK subgroup. CDPKs and SnRKs are found on all five Arabidopsis chromosomes. The presence of closely related kinases in regions of the genome known to have arisen by genome duplication indicates that these kinases probably arose by divergence from common ancestors. The PlantsP database provides a resource of continuously updated information on protein kinases from Arabidopsis and other plants.