Proteolytic processing of SERK3/BAK1 regulates plant immunity, development and cell death

Plants have evolved many receptor-like kinases (RLKs) to sense extrinsic and intrinsic cues. The signaling pathways mediated by multiple leucine-rich repeat (LRR) RLK (LRR-RLK) receptors require ligand-induced receptor-coreceptor heterodimerization and transphosphorylation with BAK1/SERK family LRR-RLKs. Here we reveal an additional layer of regulation of BAK1 via a Ca2+-dependent proteolytic cleavage process that is conserved in Arabidopsis thaliana, Nicotiana benthamiana and Saccharomyces cerevisiae . The proteolytic cleavage of BAK1 is intrinsically regulated in response to developmental cues and immune stimulation. The surface-exposed aspartic acid (D287) residue of BAK1 is critical for its proteolytic cleavage and plays an essential role in BAK1-regulated plant immunity, growth hormone brassinosteroid-mediated responses and cell death containment. BAK1D287A mutation impairs BAK1 phosphorylation on its substrate BIK1, and its plasma membrane (PM) localization. Intriguingly, it aggravates BAK1 overexpression-triggered cell death independent of BIK1, suggesting that maintaining homeostasis of BAK1 through a proteolytic process is crucial to control plant growth and immunity. Our data reveal that in addition to layered transphosphorylation in the receptor complexes, the proteolytic cleavage is an important regulatory process for the proper functions of the shared co-receptor BAK1 in diverse cellular signaling pathways

Dual-Level Regulation of ACC Synthase Activity by MPK3/MPK6 Cascade and Its Downstream WRKY Transcription Factor during Ethylene Induction in Arabidopsis

Plants under pathogen attack produce high levels of ethylene, which plays important roles in plant immunity. Previously, we reported the involvement of ACS2 and ACS6, two Type I ACS isoforms, in Botrytis cinerea–induced ethylene biosynthesis and their regulation at the protein stability level by MPK3 and MPK6, two Arabidopsis pathogen-responsive mitogen-activated protein kinases (MAPKs). The residual ethylene induction in the acs2/acs6 double mutant suggests the involvement of additional ACS isoforms. It is also known that a subset of ACS genes, including ACS6, is transcriptionally induced in plants under stress or pathogen attack. However, the importance of ACS gene activation and the regulatory mechanism(s) are not clear. In this report, we demonstrate using genetic analysis that ACS7 and ACS11, two Type III ACS isoforms, and ACS8, a Type II ACS isoform, also contribute to the B. cinerea–induced ethylene production. In addition to post-translational regulation, transcriptional activation of the ACS genes also plays a critical role in sustaining high levels of ethylene induction. Interestingly, MPK3 and MPK6 not only control the stability of ACS2 and ACS6 proteins via direct protein phosphorylation but also regulate the expression of ACS2 and ACS6 genes. WRKY33, another MPK3/MPK6 substrate, is involved in the MPK3/MPK6-induced ACS2/ACS6 gene expression based on genetic analyses. Furthermore, chromatin-immunoprecipitation assay reveals the direct binding of WRKY33 to the W-boxes in the promoters of ACS2 and ACS6 genes in vivo, suggesting that WRKY33 is directly involved in the activation of ACS2 and ACS6 expression downstream of MPK3/MPK6 cascade in response to pathogen invasion. Regulation of ACS activity by MPK3/MPK6 at both transcriptional and protein stability levels plays a key role in determining the kinetics and magnitude of ethylene induction

Phosphorylation of a WRKY Transcription Factor by Two Pathogen-Responsive MAPKs Drives Phytoalexin Biosynthesis in Arabidopsis[C][W]

WRKY33 functions downstream of pathogen-responsive MPK3 and MPK6 in reprogramming the expression of camalexin biosynthetic genes; this drives the metabolic flow to camalexin production in Arabidopsis challenged by pathogens. Biochemical and genetic analyses demonstrate that the phosphorylation of WRKY33 by MPK3/MPK6 plays an important role in the process

Phosphorylation of a WRKY Transcription Factor by MAPKs Is Required for Pollen Development and Function in <i>Arabidopsis</i>

<div><p>Plant male gametogenesis involves complex and dynamic changes in gene expression. At present, little is known about the transcription factors involved in this process and how their activities are regulated. Here, we show that a pollen-specific transcription factor, WRKY34, and its close homolog, WRKY2, are required for male gametogenesis in <i>Arabidopsis thaliana</i>. When overexpressed using <i>LAT52</i>, a strong pollen-specific promoter, epitope-tagged WRKY34 is temporally phosphorylated by MPK3 and MPK6, two mitogen-activated protein kinases (MAPKs, or MPKs), at early stages in pollen development. During pollen maturation, WRKY34 is dephosphorylated and degraded. Native promoter-driven WRKY34-YFP fusion also follows the same expression pattern at the protein level. WRKY34 functions redundantly with WRKY2 in pollen development, germination, and pollen tube growth. Loss of MPK3/MPK6 phosphorylation sites in WRKY34 compromises the function of WRKY34 <i>in vivo</i>. Epistasis interaction analysis confirmed that <i>MPK6</i> belongs to the same genetic pathway of <i>WRKY34</i> and <i>WRKY2</i>. Our study demonstrates the importance of temporal post-translational regulation of WRKY transcription factors in the control of developmental phase transitions in plants.</p></div