ORCID IDs: 0000-0002-6738-3574 (S.R.C.); 0000-0002-5570-3120 (S.P.)Signaling pathways mediated by heterotrimeric G-protein complexes comprising Ga, Gb, and Gg subunits and their regulatory RGS (Regulator of G-protein Signaling) protein are conserved in all eukaryotes. We have shown that the specific Gb and Gg proteins of a soybean (Glycine max) heterotrimeric G-protein complex are involved in regulation of nodulation. We now demonstrate the role of Nod factor receptor 1 (NFR1)-mediated phosphorylation in regulation of the G-protein cycle during nodulation in soybean. We also show that during nodulation, the G-protein cycle is regulated by the activity of RGS proteins. Lower or higher expression of RGS proteins results in fewer or more nodules, respectively. NFR1 interacts with RGS proteins and phosphorylates them. Analysis of phosphorylated RGS protein identifies specific amino acids that, when phosphorylated, result in significantly higher GTPase accelerating activity. These data point to phosphorylation-based regulation of G-protein signaling during nodule development. We propose that active NFR1 receptors phosphorylate and activate RGS proteins, which help maintain the Ga proteins in their inactive, trimeric conformation, resulting in successful nodule development. Alternatively, RGS proteins might also have a direct role in regulating nodulation because overexpression of their phospho-mimic version leads to partial restoration of nodule formation in nod49 mutants.

Receptors form the crux for any biochemical signaling. Receptor-like kinases (RLKs) are conserved protein kinases in eukaryotes that establish signaling circuits to transduce information from outer plant cell membrane to the nucleus of plant cells, eventually activating processes directing growth, development, stress responses, and disease resistance. Plant RLKs share considerable homology with the receptor tyrosine kinases (RTKs) of the animal system, differing at the site of phosphorylation. Typically, RLKs have a membrane-localization signal in the amino-terminal, followed by an extracellular ligand-binding domain, a solitary membrane-spanning domain, and a cytoplasmic kinase domain. The functional characterization of ligand-binding domains of the various RLKs has demonstrated their essential role in the perception of extracellular stimuli, while its cytosolic kinase domain is usually confined to the phosphorylation of their substrates to control downstream regulatory machinery. Identification of the several ligands of RLKs, as well as a few of its immediate substrates have predominantly contributed to a better understanding of the fundamental signaling mechanisms. In the model plant Arabidopsis, several studies have indicated that multiple RLKs are involved in modulating various types of physiological roles via diverse signaling routes. Here, we summarize recent advances and provide an updated overview of transmembrane RLKs in Arabidopsis.

Plant defense responses at stomata and apoplast are the most important early events during plant–bacteria interactions. The key components of stomatal defense responses have not been fully characterized. A GTPase encoding gene, NOG1-2, which is required for stomatal innate immunity against bacterial pathogens, was recently identified. Functional studies in Arabidopsis revealed that NOG1-2 regulates guard cell signaling in response to biotic and abiotic stimulus through jasmonic acid (JA)- and abscisic acid (ABA)-mediated pathways. Interestingly, in this study, Jasmonate-ZIM-domain protein 9 (JAZ9) was identified to interact with NOG1-2 for the regulation of stomatal closure. Upon interaction, JAZ9 reduces GTPase activity of NOG1-2. We explored the role of NOG1-2 binding with JAZ9 for COI1-mediated JA signaling and hypothesized that its function may be closely linked to MYC2 transcription factor in the regulation of the JA-signaling cascade in stomatal defense against bacterial pathogens. Our study provides valuable information on the function of a small GTPase, NOG1-2, in guard cell signaling and early plant defense in response to bacterial pathogens.

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Heterotrimeric G-proteins influence almost all aspects of plant growth, development, and responses to biotic and abiotic stresses in plants, likely via their interaction with specific effectors. However, the identity of such effectors and their mechanism of action are mostly unknown. While investigating the roles of different G-protein subunits in modulating the oil content in Camelina (Camelina sativa), an oil seed crop, we uncovered a role of Gb proteins in controlling anisotropic cell expansion. Knockdown of Gb genes causes reduced longitudinal and enhanced transverse expansion, resulting in altered cell, tissue, and organ shapes in transgenic plants during vegetative and reproductive development. These plants also exhibited substantial changes in their fatty acid and phospholipid profiles, which possibly leads to the increased oil content of the transgenic seeds. This increase is potentially caused by the direct interaction of Gb proteins with a specific patatin-like phospholipase, pPLAIIId. Camelina plants with suppressed Gb expression exhibit higher lipase activity, and show phenotypes similar to plants overexpressing pPLAIIId, suggesting that the Gb proteins are negative regulators of pPLAIIId. These results reveal interactions between the G-protein-mediated and lipid signaling/metabolic pathways, where specific phospholipases may act as effectors that control key developmental and environmental responses of plants.

Drought is the most prevalent unfavorable condition that impairs plant growth and development by altering morphological, physiological, and biochemical functions, thereby impeding plant biomass production. To survive the adverse effects, water limiting condition triggers a sophisticated adjustment mechanism orchestrated mainly by hormones that directly protect plants via the stimulation of several signaling cascades. Predominantly, water deficit signals cause the increase in the level of endogenous ABA, which elicits signaling pathways involving transcription factors that enhance resistance mechanisms to combat drought-stimulated damage in plants. These responses mainly include stomatal closure, seed dormancy, cuticular wax deposition, leaf senescence, and alteration of the shoot and root growth. Unraveling how plants adjust to drought could provide valuable information, and a comprehensive understanding of the resistance mechanisms will help researchers design ways to improve crop performance under water limiting conditions. This review deals with the past and recent updates of ABA-mediated molecular mechanisms that plants can implement to cope with the challenges of drought stress.

Background Cyclin-dependent kinases (CDKs) are a predominant group of serine/threonine protein kinases that have multi-faceted functions in eukaryotes. The plant CDK members have well-known roles in cell cycle progression, transcriptional regulation, DNA repair, abiotic stress and defense responses, making them promising targets for developing stress adaptable high-yielding crops. There is relatively sparse information available on the CDK family genes of cultivated oilseed crop peanut and its diploid progenitors. Results We have identified 52 putative cyclin-dependent kinases (CDKs) and CDK-like (CDKLs) genes in Arachis hypogaea (cultivated peanut) and total 26 genes in each diploid parent of cultivated peanut (Arachis duranensis and Arachis ipaensis). Both CDK and CDKL genes were classified into eight groups based on their cyclin binding motifs and their phylogenetic relationship with Arabidopsis counterparts. Genes in the same subgroup displayed similar exon–intron structure and conserved motifs. Further, gene duplication analysis suggested that segmental duplication events played major roles in the expansion and evolution of CDK and CDKL genes in cultivated peanuts. Identification of diverse cis-acting response elements in CDK and CDKL genes promoter indicated their potential fundamental roles in multiple biological processes. Various gene expression patterns of CDKs and CDKLs in different peanut tissues suggested their involvement during growth and development. In addition, qRT-PCR analysis demonstrated that most representing CDK and CDKL gene family members were significantly down-regulated under ABA, PEG and mannitol treatments. Conclusions Genome-wide analysis offers a comprehensive understanding of the classification, evolution, gene structure, and gene expression profiles of CDK and CDKL genes in cultivated peanut and their diploid progenitors. Additionally, it also provides cell cycle regulatory gene resources for further functional characterization to enhance growth, development and abiotic stress tolerance.

1 Heterotrimeric G-proteins, comprised of Gα, Gβ and Gγ subunits regulate signaling in 2 eukaryotes. In metazoans, G-proteins are activated by GPCR-mediated GDP to GTP 3 exchange on Gα; however, the role of receptors in regulating plant G-protein signaling 4 remains equivocal. Mounting evidence points to the involvement of receptor-like kinases 5 (RLKs) in regulating plant G-protein signaling pathways, but their mechanistic details 6 remain limited. We have previously shown that during soybean nodulation, the nod factor 7 receptor 1 (NFR1) interacts with G-protein components and indirectly controls signaling. 8  We explored the direct regulation of G-protein signaling by RLKs using protein-protein 9 interactions, receptor-mediated phosphorylation and the effects of such phosphorylations 10 on soybean nodule formation. 11 Results presented in this study demonstrate a direct, phosphorylation-based regulation of 12 Gα by symbiosis receptor kinase (SymRK). SymRKs interact with and phosphorylate Gα 13 at multiple residues, including two in its active site, which abolishes GTP binding. In 14 addition, phospho-mimetic Gα fails to interact with Gβγ, potentially allowing for 15 constitutive signaling by the freed Gβγ. 16 These results uncover a novel mechanism of G-protein cycle regulation in plants where 17 receptor-mediated phosphorylation of Gα not only affects its activity, but also influences 18 the availability of its signaling partners, thereby exerting a two-pronged control on 19 signaling. 20 100 Intriguingly, two of the phosphorylated amino acids are in the conserved GTP-binding domain of 101 the Gα protein. Phosphorylation of the serine residues in this site abolishes the ability of Gα to 102 bind GTP, essentially making the protein biochemically inactive. Furthermore, the phospho-103 mimetic Gα is unable to bind Gβγ proteins, suggesting an altered dynamics and/or availability of 104 the active proteins for downstream signaling. Our results thus uncover a novel G-protein signaling 105 mechanism where not only the activity but also the availability of the signaling proteins is 106 dependent on receptor-mediated phosphorylation of the Gα protein. 107 MATERIAL AND METHODS 108 Plant material and hairy root transformation 109 Soybean (Glycine max) wild type ('Williams 82') seeds were grown on (BM 7 35%) in the 110 greenhouse (16 h light/8 h dark) for 12 days at 25°C. Hairy root transformation was performed as Rhizobium strain (USDA136) cultured in Vincent's rich medium was used for bacterial infection. 113 Nodules were counted at 32 days after rhizobium infection. Three biological replicates were used 114 for each construct. At least 35 to 40 transgenic hairy roots were used in individual experiment for 115 each construct. The data were averaged, and statistically significant values were determined by 116 Dunn's multiple comparisons test. 117 Generation of constructs 118 The gene fragment for RNAi construct of SymRK was cloned into the pCR8/GW vector 119 (Invitrogen) and subsequently transferred into binar...