Heterotrimeric G protein signalling in the plant kingdom

In animals, heterotrimeric G proteins, comprising α-, β-and γ-subunits, perceive extracellular stimuli through cell surface receptors, and transmit signals to ion channels, enzymes and other effector proteins to affect numerous cellular behaviours. In plants, G proteins have structural similarities to the corresponding molecules in animals but transmit signals by atypical mechanisms and effector proteins to control growth, cell proliferation, defence, stomate movements, channel regulation, sugar sensing and some hormonal responses. In this review, we summarize the current knowledge on the molecular regulation of plant G proteins, their effectors and the physiological functions studied mainly in two model organisms: Arabidopsis thaliana and rice (Oryza sativa). We also look at recent progress on structural analyses, systems biology and evolutionary studies.     

异三聚体G蛋白在植物非生物胁迫应答中的作用

异三聚体G蛋白(Heterotrimeric GTP-binding proteins)是真核生物中一类重要的信号传导分子,由Gα、Gβ和Gγ3个亚基组成。异三聚体G蛋白不仅参与了植物的生长发育调控,而且还在多种非生物胁迫应答中起着重要的调控作用。本文着重介绍异三聚体G蛋白在植物非生物胁迫应答中的作用及可能的调控机制,并结合当前研究现状对未来研究方向提出展望,以期为今后深入研究异三聚体G蛋白在植物非生物胁迫应答中的调控机制提供参考。     

Do plants have rhodopsin after all? A mystery of plant G protein-coupled signalling

Considerable physiological and biochemical evidence suggests that plants, like animals, widely use intracellular signalling coupled to heterotrimeric G proteins. Yet, the molecular components of this machinery remained elusive until recently. We overview the work carried out during the last two decades, aimed at identification of the plant proteins involved in G protein-coupled signalling. The completion of the sequencing of the Arabidopsis genome now permits to assess whether plants possess signalling and regulatory components of this machinery corresponding to those known from animals.     

植物G蛋白与植物防卫反应

近年来, 植物G蛋白(包括异三聚体G蛋白和小G蛋白)的存在及其信号调控途径已经成为人们研究细胞信号转导过程的热点问题。从多种植物细胞中相继分离克隆出多个与动物G蛋白同源的编码植物G蛋白的基因, 并且植物G蛋白的种类和数量有其独特性。植物G蛋白在植物细胞跨膜信号转导中发挥重要的作用, 参与多种生命活动的调控。本文主要综述了植物G蛋白参与植物防卫反应调节作用的研究进展。     

Small GTP-binding Proteins and their Functions in Plants

Small GTP-binding proteins exist in eukaryotes from yeast to animals to plants and constitute a superfamily whose members function as molecular switches that cycle between “active” and “inactive” states. They regulate a wide variety of cell functions such as signal transduction, cell proliferation, cytoskeletal organization, intracellular membrane trafficking, and gene expression. In yeast and animals, this superfamily is structurally classified into at least five families: the Ras, Rho, Rab, Arf/Sar1, and Ran families. However, plants contain Rab, Rho, Arf, and Ran homologs, but no Ras. Small GTP-binding proteins have become an intensively studied group of regulators not only in yeast and animals but also in plants in recent years. In this article we briefly review the class and structure of small GTP-binding proteins. Their working modes and functions in animals and yeast are listed, and the functions of individual members of these families in plants are discussed, with the emphasis on the recently revealed plant-specific roles of these proteins, including their cross-talk with plant hormones and other signals, regulation of organogenesis (leaf, root, and embryo), polar growth, cell division, and involvement in various stress and defense responses.     

The Rop GTPase: an emerging signaling switch in plants

G proteins are ubiquitous molecular switches in eukaryotic signal transduction, but their roles in plant signal transduction had not been clearly established until recent studies of the plant-specific Rop subfamily of RHO GTPases. Rop participates in signaling to an array of physiological processes including cell polarity establishment, cell growth, morphogenesis, actin dynamics, H₂O₂ generation, hormone responses, and probably many other cellular processes in plants. Evidence suggests that plants have developed unique molecular mechanisms to control this universal molecular switch through novel GTPase-activating proteins and potentially through a predominant class of plant receptor-like serine/threonine kinases. Furthermore, the mechanism by which Rop regulates specific processes may also be distinct from that for other GTPases. These advances have raised the exciting possibility that the elucidation of Rop GTPase signaling may lead to the establishment of a new paradigm for G protein-dependent signal transduction in plants.     

Regulation of root-wave response by extra large and conventional G proteins in Arabidopsis thaliana

Heterotrimeric G proteins composed of α, β and γ subunits regulate a number of fundamental processes concerned with growth and development in plants. In addition to the canonical heterotrimeric G proteins, plants also contain a small family of extra large G proteins (XLGs) that show significant similarity to the G-protein α subunit in their C-terminal regions. In this paper we show that one of the three XLG genes, XLG3 , and the Gβ subunit (AGB1) of the Arabidopsis G-protein heterotrimer are specifically involved in the regulation of a subset of root morphological and growth responses. Based on analysis of T-DNA insertional mutant phenotypes, XLG3 and AGB1 each positively regulate root waving and root skewing. Since these responses are regulated by physical as well as physiological cues, we assessed the roles of AGB1 and XLG3 in gravitropism, thigmotropism and hormonal responses. Our data show that mutants lacking either XLG3 or AGB1 genes are hypersensitive to ethylene and show growth responses consistent with alterations in auxin transport, while maintaining an essentially wild-type response to the physical cues of gravity and touch. These results suggest that XLG3 and AGB1 proteins regulate the hormonal determinants of root-waving and root-skewing responses in plants and possibly interact in a tissue-specific or signal-specific manner. Because plants harboring knockout mutations in the Gα subunit gene, GPA1 , exhibit wild-type root waving and skewing, our results may indicate that the AGB1 subunit functions in these processes without formation of a classic Gαβγ heterotrimer.     

植物G蛋白偶联受体及其在信号转导中的作用

G蛋白偶联受体(G protein-coupled receptors,GPCRs)是具有7个跨膜螺旋的蛋白质受体,是人体内最大的蛋白质超家族.GPCRs能调控细胞周期,参与多种植物信号通路以及影响一系列的代谢和分化活动.简要介绍了GPCR和G蛋白介导的信号转导机制,GPCRs的结构和植物GPCR及其在植物跨膜信号转导中的作用,并对GPCR的信号转导机制及植物抗病反应分子机制的研究提出展望.     

Flexible functional interactions between G‐protein subunits contribute to the specificity of plant responses

Plants being sessile integrate information from a variety of endogenous and external cues simultaneously to optimize growth and development. This necessitates the signaling networks in plants to be highly dynamic and flexible. One such network involves heterotrimeric G‐proteins comprised of Gα, Gβ, and Gγ subunits, which influence many aspects of growth, development, and stress response pathways. In plants such as Arabidopsis, a relatively simple repertoire of G‐proteins comprised of one canonical and three extra‐large Gα, one Gβ and three Gγ subunits exists. Because the Gβ and Gγ proteins form obligate dimers, the phenotypes of plants lacking the sole Gβ or all Gγ genes are similar, as expected. However, Gα proteins can exist either as monomers or in a complex with Gβγ, and the details of combinatorial genetic and physiological interactions of different Gα proteins with the sole Gβ remain unexplored. To evaluate such flexible, signal‐dependent interactions and their contribution toward eliciting a specific response, we have generated Arabidopsis mutants lacking specific combinations of Gα and Gβ genes, performed extensive phenotypic analysis, and evaluated the results in the context of subunit usage and interaction specificity. Our data show that multiple mechanistic modes, and in some cases complex epistatic relationships, exist depending on the signal‐dependent interactions between the Gα and Gβ proteins. This suggests that, despite their limited numbers, the inherent flexibility of plant G‐protein networks provides for the adaptability needed to survive under continuously changing environments.     

丝瓜的三聚体G蛋白的初步研究

根据拟南芥（ａｒａｂｉｄｏｐｓｉｓｔｈａｈｌｉａｎａ）ＧＰＡ１的保守区段Ａ设计一对特异引物（５′ｃｔｇｇｇｇａａｔｃｔｇｇａａａａｔｃ３′,５′ｃａｃａｇｃｔｇｔａｃａｃｃｔｃａａａｃ３′）通过ＰＣＲ从丝瓜核基因组中扩增植物的三聚体Ｇ蛋白α亚基编码基因,获得了２个片段（ＬＦＧ１,ＬＦＧ２）,并已克隆和测序（已在ＥＭＢＬ数据库中登记,登记号为：ｙ１５２７０,ｙ１５２７１）．序列分析表明ＬＦＧ１和ＬＦＧ２分别由１５１５ｂｐ和７３２ｂｐ构成,都含有三聚体Ｇ蛋白α亚基编码基因的保守区段Ａ,但也都含有内含子．根据片段的大小及ＰＣＲ的特性,ＬＦＧ１可能是丝瓜三聚体Ｇ蛋白α亚基编码基因上的片段．     

植物胞外钙调素与信号转导

植物体需要构建复杂的信号转导体系以调节自身的生长发育过程并适应外界环境的变化,这种功能的实现需要胞内和胞外诸多信号分子的参与,胞外钙调素的发现使人们开始相信植物细胞外多肽信使的存在。胞外钙调素的生物学功能极其广泛,几乎涉及到植物生长发育的各个阶段,其信号转导途径是目前研究得最多也是最为清楚的方面,异三聚体G蛋白、磷脂酶C(PLC)-肌醇三磷酸(IP3)-肌醇三磷酸受体(IP3R)信号通路、活性氧和Ca2 通道之间直接或间接的相互作用是胞外钙调素信号转导的核心。     

Serpentine type receptors and heterotrimeric G-proteins in yeasts: Structural-functional organization and molecular mechanisms of action

The signal systems of the yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe, coupled to heterotrimeric G-proteins and sensitive to pheromones and alimentary molecules, are prototypes of hormonal signal systems of the higher vertebrate animals and are widely used in studies on molecular mechanisms of their functioning. This review summarizes and analyzes data on structural-functional organization of the first two components of these systems—receptors of the serpentine type and heterotrimeric G-proteins; mechanisms of functional coupling of receptors and G-proteins both between each other and to other signal proteins are discussed. It has been shown that at the early stages of evolution of signaling systems, at the yeast level, various models of transduction of signals into the cell were tested; many of them differ essentially from the classic model of the three-component, G-protein-coupled signal system of the higher vertebrates.     

“Round Up the Usual Suspects”: A Comment on Nonexistent Plant G Protein-Coupled Receptors

An evolutionary argument supports the conclusion that plants do not have G protein coupled receptors.In the classic 1942 movie Casablanca, Vichy Police Captain Louis Renault obfuscated the truth by commanding his lieutenants to “round up the usual suspects,” knowing well that the culprit with the gun stood in plain view. Something similar has happened in the plant G protein field. This Scientific Correspondence was written to shed light on the source of misunderstanding and to preempt further confusion. Plant heterotrimeric G proteins are self-activating and therefore do not need and do not utilize G protein-coupled receptors (GPCRs). This conclusion was reached previously from biochemical analyses of plant G proteins (Johnston et al., 2007a; Urano et al., 2012); here, we buttress this point of view using an evolutionary argument. Proteins suspected as plant GPCRs were “rounded up” because they have the predicted topology of animal GPCRs and/or have been misannotated as such; however, these proteins are highly conserved in organisms that lack heterotrimeric G proteins. Therefore, they have functions unrelated to G-coupled signaling. Instead, the culprit protein standing in plain view is a receptor GTPase-accelerating protein (GAP), a receptor GAP called AtRGS1 (for regulator of G signaling).     

Regulation of vesicular neurotransmitter transporters

Neurotransmitters are key molecules of neurotransmission. They are concentrated first in the cytosol and then in small synaptic vesicles of presynaptic terminals by the activity of specific neurotransmitter transporters of the plasma and the vesicular membrane, respectively. It has been shown that postsynaptic responses to single neurotransmitter packets vary over a wide range, which may be due to a regulation of vesicular neurotransmitter filling. Vesicular filling depends on the availability of transmitter molecules in the cytoplasm and the active transport into secretory vesicles relying on a proton gradient. In addition, it is modulated by vesicle-associated heterotrimeric G proteins, Go2 and Gq, which regulate VMAT activities in brain and platelets, respectively, and may also be involved in the regulation of VGLUTs. It appears that the vesicular content activates the G protein, suggesting a signal transduction form the luminal site which might be mediated by a vesicular G-protein coupled receptor or, as an alternative, possibly by the transporter itself. These novel functions of G proteins in the control of transmitter storage may link regulation of the vesicular content to intracellular signal cascades.     

The stimulation of NADH oxidase activity of rat liver plasma membranes by guanine nucleotides may involve both guanine nucleotide-binding proteins of the plasma membrane and responses not mediated by classic heterotrimeric G proteins

Summary The stimulation of NADH oxidase activity of plasma membranes of rat liver observed with guanine nucleotides may involve both guanine nucleotide-binding proteins of the plasma membrane and responses not mediated by classic heterotrimeric G proteins. These conclusions are based on findings that detergent treatment and peptide antisera to a consensus guanine nucleotidebinding domain (GAGES) of G subunits of heterotrimeric G proteins reduced but did not eliminate the stimulation of NADH oxidase activity by guanine nucleotides. The proteins immunoprecipitated by the antisera, when added back to plasma membranes, stimulated the NADH oxidase activity. This stimulated rate was further stimulated by the addition of GTP but was not dependent upon guanine nucleotide presence. Additions of cytosol, either fractionated or unfractionated did not appear to stimulate the NADH oxidase activity of rat liver plasma membranes. The activities of the plasma membranes and the activities introduced by the cytosol fractions were nearly, but not entirely, additive. The results are suggestive of a subunit composition of the NADH oxidase but one distinct from that involving solely heterotrimeric G proteins. Also a strong dependence on cytosolic components, as found with the NADPH oxidase complex of neutrophils, is not obvious. In addition, the possibility that the NADH oxidase may exhibit an intrinsic re-sponse to guanine nucleotides, not dependent on accessory proteins, cannot be ruled out. Among the several bands immunoprecipitated with the antisera and reactive with the antisera on Western blots, were peptide bands in the molecular weight range ascribed to the NADH oxidase.     

Activators of G proteins inhibit GSK-3β and stabilize β-Catenin in Xenopus oocytes

Frizzled proteins, the receptors for Wnt ligands have seven hydrophobic transmembrane domains, a structural feature of G protein coupled receptors. Therefore a role for G proteins in the regulation of Wnt signaling has been proposed. Here I have used Xenopus oocytes to study the role of heterotrimeric G proteins in the regulation of GSK-3β and β-Catenin, two essential components of the canonical Wnt pathway. In these cells, general activators of G proteins such as GTPγ-S and AlF4⁻ increase β-Catenin stability and decrease GSK-3β mediated phosphorylation of the microtubule associated protein, Tau. Among several members of Gα proteins tested, expression of a constitutively active mutant of Gαq (GαqQL) led to a significant increase in accumulation of β-Catenin. The stabilization of β-Catenin mediated by Gαq was reversed by a Gαq specific inhibitor, Gp-antagonist 2A, but not by a specific blocking peptide for Gαs. Expression of GαqQL also inhibited GSK-3β-mediated tau phosphorylation in Xenopus oocytes. These results support a role for the Gq class of G proteins in the regulation of Wnt/β-Catenin signal transduction.     

Arabidopsis extra-large G proteins (XLGs) regulate root morphogenesis

As in mammalian systems, heterotrimeric G proteins, composed of alpha, beta and gamma subunits, are present in plants and are involved in the regulation of development and cell signaling. Besides the sole prototypical G protein alpha subunit gene, GPA1, the Arabidopsis thaliana genome has three extra-large GTP-binding protein (XLG)-encoding genes: XLG1 (At2g23460), XLG2 (At4g34390) and XLG3 (At1g31930). The C-termini of the XLGs are Galpha domains that are homologous to GPA1, whereas their N-termini each contain a cysteine-rich region and a putative nuclear localization signal (NLS). GFP fusions with each XLG confirmed nuclear localization. All three XLG genes are expressed in essentially all plant organs, with strong expression in vascular tissues, primary root meristems and lateral root primordia. Analysis of single, double and triple T-DNA insertional mutants of the XLG genes revealed redundancy in XLG function. Dark-grown xlg1-1 xlg2-1 xlg3-1 triple mutant plants showed markedly increased primary root length compared with wild-type plants. This phenotype was not observed in dark-grown xlg single mutants, and was suppressed upon complementation of the xlg triple mutant with each XLG. Root cell sizes of the xlg triple mutant and root morphology were highly similar to those of wild-type roots, suggesting that XLGs may regulate cell proliferation. Dark-grown roots of the xlg triple mutants also showed altered sensitivity to sugars, ABA hyposensitivity and ethylene hypersensitivity, whereas seed germination in xlg triple mutants was hypersensitive to osmotic stress and ABA. As plant-specific proteins, regulatory mechanisms of XLGs may differ from those of conventional Galphas.     

Signal transduction by plant heterotrimeric G‐protein

Heterotrimeric G‐proteins are complexes that regulate important signalling pathways essential for growth and development in both plants and animals. Although plant cells are composed of the core components (Gα, Gβ and Gγ subunits) found in animal G‐proteins, the complexities of the architecture, function and signalling mechanisms of those in animals are dissimilar to those identified in some plants. Current studies on plant G‐proteins have improved knowledge of the essential physiological and agronomic properties, which when harnessed, could potentially impact global food security. Extensive studies on the molecular mechanisms underlying these properties in diverse plant species will be imperative in improving our current understanding of G‐protein signalling pathways involved in plant growth and development. The advancement of G‐protein signalling networks in distinct plant species could significantly aid in better crop development. This review summarizes current progress, novel discoveries and future prospects for this area in potential crop improvement.     

Function of Heterotrimeric G Protein in Gibberellin Signaling

The importance of plant heterotrimeric G protein functions has recently been recognized. Rice and Arabidopsis mutants of genes coding the subunits of the G proteins have been isolated and physiological studies on these mutants have suggested that plant heterotrimeric G proteins are involved in several intra-signaling pathways driven by external signals, such as gibberellin, auxin, abscisic acid, brassinolide, ethylene, light, and elicitor. The possible functions of rice heterotrimeric G proteins in gibberellin signaling are discussed here.     

Structure and function of heterotrimeric G proteins in plants

Heterotrimeric G proteins are mediators that transmit the external signals via receptor molecules to effector molecules. The G proteins consist of three different subunits: alpha, beta, and gamma subunits. The cDNAs or genes for all the alpha, beta, and gamma subunits have been isolated from many plant species, which has contributed to great progress in the study of the structure and function of the G proteins in plants. In addition, rice plants lacking the alpha subunit were generated by the antisense method and a rice mutant, Daikoku d1, was found to have mutation in the alpha-subunit gene. Both plants show abnormal morphology such as dwarfism, dark green leaf, and small round seed. The findings revealed that the G proteins are functional molecules regulating some body plans in plants. There is evidence that the plant G proteins participate at least in signaling of gibberellin at low concentrations. In this review, we summarize the currently known information on the structure of plant heterotrimeric G proteins and discuss the possible functions of the G proteins in plants.