Plants deploy numerous cell surface-localized pattern-recognition receptors (PRRs) to perceive host- and microbe-derived molecular patterns that are specifically released during infection and activate defense responses. The activation of the mitogen-activated protein kinases MPK3, MPK4, and MPK6 (MPK3/4/6) is a hallmark of immune system activation by all known PRRs and is crucial for establishing disease resistance. The MAP kinase kinase kinase (MAPKKK) MEKK1 controls MPK4 activation, but the MAPKKKs responsible for MPK3/6 activation downstream of diverse PRRs and how the perception of diverse molecular patterns leads to the activation of MAPKKKs remain elusive. Here, we show that two highly related MAPKKKs, MAPKKK3 and MAPKKK5, mediate MPK3/6 activation by at least four PRRs and confer resistance to bacterial and fungal pathogens in Arabidopsis thaliana. The receptor-like cytoplasmic kinases VII (RLCK VII), which act downstream of PRRs, directly phosphorylate MAPKKK5 Ser-599, which is required for pattern-triggered MPK3/6 activation, defense gene expression, and disease resistance. Surprisingly, MPK6 further phosphorylates MAPKKK5 Ser-682 and Ser-692 to enhance MPK3/6 activation and disease resistance, pointing to a positive feedback mechanism. Finally, MEKK1 Ser-603 is phosphorylated by both RLCK VII and MPK4, which is required for pattern-triggered MPK4 activation. These findings illustrate central mechanisms by which multiple PRRs activate MAPK cascades and disease resistance.

Pattern-recognition receptors (PRRs), which consist of receptor kinases (RKs) and receptor-like proteins (RLPs), sense microbial- and host-derived molecular patterns associated with pathogen infection to trigger immune responses in plants. Several kinases of the 46-member Arabidopsis thaliana receptor-like cytoplasmic kinase (RLCK) subfamily VII play important roles in pattern-triggered immunity, but it is unclear whether different RLCK VII members act specifically or redundantly in immune signaling. Here, we constructed nine higher order mutants of this subfamily (named rlck vii-1 to rlck vii-9) and systematically characterized their immune phenotypes. The mutants rlck vii-5, -7, and -8 had compromised reactive oxygen species (ROS) production in response to all patterns tested, indicating that the corresponding members are broadly required for signaling of multiple PRRs. However, rlck vii-4 was specifically defective in chitin-induced ROS production, suggesting that RCLK VII-4 members mediate the signaling of specific PRRs. Furthermore, RLCK VII-4 members were required for chitin-triggered activation of mitogen-activated protein kinase (MAPK), demonstrating that these kinases link a PRR to MAPK activation. Moreover, we found that RLCK VII-6 and -8 were also required for RK-mediated root growth. Together these results show that numerous RLCK VII members are involved in pattern-triggered immune signaling and uncover both common and specific roles of these kinases in plant development and immunity mediated by various RKs.

Fruit length is a key domestication trait that affects crop yield and appearance. Cucumber (Cucumis sativus) fruits vary from 5 to 60 cm in length. Despite the identification of several regulators and multiple quantitative trait loci (QTLs) underlying fruit length, the natural variation, and molecular mechanisms underlying differences in fruit length are poorly understood. Through map-based cloning, we identified a nonsynonymous polymorphism (G to A) in CRABS CLAW (CsCRC) as underlying the major-effect fruit size/shape QTL FS5.2 in cucumber. The short-fruit allele CsCRC^A is a rare allele that has only been found in round-fruited semi-wild Xishuangbanna cucumbers. A near-isogenic line (NIL) homozygous for CsCRC^A exhibited a 34∼39% reduction in fruit length. Introducing CsCRC^G into this NIL rescued the short-fruit phenotype, and knockdown of CsCRC^G resulted in shorter fruit and smaller cells. In natural cucumber populations, CsCRC^G expression was positively correlated with fruit length. Further, CsCRC^G, but not CsCRC^A, targets the downstream auxin-responsive protein gene CsARP1 to regulate its expression. Knockout of CsARP1 produced shorter fruit with smaller cells. Hence, our work suggests that CsCRC^G positively regulates fruit elongation through transcriptional activation of CsARP1 and thus enhances cell expansion. Using different CsCRC alleles provides a strategy to manipulate fruit length in cucumber breeding.

WUSCHEL-related homeobox (WOX) transcription factors are plant-specific members that characterized by the presence of a homeodomain. It has been shown that WOX members regulate several aspects of plant development, but the biological functions of this CsWOX gene family remain largely unknown in cucumber (Cucumis sativus L.). In this study, we identified and characterized 11 putative CsWOX genes in cucumber, which are also divided into three major clades (e.g., the Ancient clade, the Intermediate clade and the WUS clade). Expression pattern analysis revealed tissue-specific expression patterns of CsWOX genes, including that CsWOX9 is mainly expressed in developing fruit and also has lower expression in tip and axillary bud, which was further confirmed by in situ hybridization assay. Moreover, overexpression of CsWOX9 in Arabidopsis led to increased branches and rosette leaves, and shorter siliques. Together, these results indicated that CsWOX members may regulate cucumber growth and development.