Protein kinases regulate virtually all cellular processes, but it remains challenging to determine the functions of all protein kinases, collectively called the “kinome”, in any species. We developed a computational approach called EXPLICIT-Kinase to predict the functions of the Arabidopsis kinome. Because the activities of many kinases can be regulated transcriptionally, their gene expression patterns provide clues to their functions. A universal gene expression predictor for Arabidopsis was constructed to predict the expression of 30,172 non-kinase genes based on the expression of 994 kinases. The model reconstituted highly accurate transcriptomes for diverse Arabidopsis samples. It identified the significant kinases as predictor kinases for predicting the expression of Arabidopsis genes and pathways. Strikingly, these predictor kinases were often regulators of related pathways, as exemplified by those involved in cytokinesis, tissue development, and stress responses. Comparative analyses revealed that portions of these predictor kinases are shared and conserved between Arabidopsis and maize. As an example, we identified a conserved predictor kinase, RAF6, from a stomatal movement module. We verified that RAF6 regulates stomatal closure. It can directly interact with SLAC1, a key anion channel for stomatal closure, and modulate its channel activity. Our approach enables a systematic dissection of the functions of the Arabidopsis kinome.

Heterotrimeric G proteins consisting of Gα, Gβ and Gγ are conserved signaling hubs in eukaryotes. Without analogs to canonical animal G protein‐coupled receptors, plant cells are thought to use RGS1 and a yet unknown mechanism to regulate the activity of Gα. Meanwhile, the exact role of canonical Gα in plant innate immunity remains controversial. Here, we report multiple immune deficiencies in the null allele of Arabidopsis Gα (GPA1) in response to bacterial flg22 elicitor, clarifying a positive regulatory role of GPA1 in flg22 signaling. We also detect overall increased phosphorylation of GPA1 but reduced phosphorylation at Thr¹⁹ upon flg22 elicitation. Interestingly, flg22 could not induce phosphorylation of GPA1^T19A and GPA1^T19D, suggesting that the dynamic Thr¹⁹ phosphorylation is required for GPA1 to respond to flg22. Moreover, flg22‐induced GPA1 phosphorylation is largely abolished in the absence of BAK1 in vivo, and BAK1 could phosphorylate GPA1 but not GPA1^T19A in vitro at the phosphorylation sites identified in vivo, suggesting BAK1 is likely the kinase for GPA1 phosphorylation in response to flg22. Furthermore, the T19A mutation could promote flg22‐induced association, rather than dissociation, between GPA1 and RGS1. Taken together, our findings shed new insights into the function and regulation of GPA1 in Arabidopsis defense signaling.

Brassinosteroids (BR) are involved in the control of several developmental processes ranging from root elongation to senescence and adaptation to environmental cues. Thus, BR perception and signaling have to be precisely regulated. One regulator is BRI1‐associated kinase 1 (BAK1)‐interacting receptor‐like kinase 3 (BIR3). In the absence of BR, BIR3 forms complexes with BR insensitive 1 (BRI1) and BAK1. However, the biophysical and energetic requirements for complex formation in the absence of the ligand have yet to be determined. Using computational modeling, we simulated the potential complexes between the cytoplasmic domains of BAK1, BRI1 and BIR3. Our calculations and experimental data confirm the interaction of BIR3 with BAK1 and BRI1, with the BAK1 BIR3 interaction clearly favored. Furthermore, we demonstrate that BIR3 and BRI1 share the same interaction site with BAK1. This suggests a competition between BIR3 and BRI1 for binding to BAK1, which results in preferential binding of BIR3 to BAK1 in the absence of the ligand thereby preventing the active participation of BAK1 in BR signaling. Our model also suggests that BAK1 and BRI1 can interact even while BAK1 is in complex with BIR3 at an additional binding site of BAK1 that does not allow active BR signaling.

Post-translational modifications of proteins by small ubiquitin-like modifiers (SUMOs) play crucial roles in plant growth and development, and in stress responses. The MMS21 is a newly-identified Arabidopsis thaliana L. SUMO E3 ligase gene aside from the SIZ1, and its function requires further elucidation. Here, we show that MMS21 deficient plants display improved drought tolerance, and constitutive expression of MMS21 reduces drought tolerance. The expression of MMS21 was reduced by abscisic acid (ABA), polyethylene glycol (PEG) or drought stress. Under drought conditions, mms21 mutants showed the highest survival rate and the slowest water loss, and accumulated a higher level of free proline compared to wild-type (WT) and MMS21 over-expression plants. Stomatal aperture, seed germination and cotyledon greening analysis indicated that mms21 was hypersensitive to ABA. Molecular genetic analysis revealed that MMS21 deficiency led to elevated expression of a series of ABA-mediated stress-responsive genes, including COR15A, RD22, and P5CS1 The ABA and drought-induced stress-responsive genes, including RAB18, RD29A and RD29B, were inhibited by constitutive expression of MMS21. Moreover, ABA-induced accumulation of SUMO-protein conjugates was blocked in the mms21 mutant. We thus conclude that MMS21 plays a role in the drought stress response, likely through regulation of gene expression in an ABA-dependent pathway.