Stomatal Development and Patterning Are Regulated by Environmentally Responsive Mitogen-Activated Protein Kinases inArabidopsis

Abstract

Stomata are specialized epidermal structures that regulate gas (CO 2 and O 2 ) and water vapor exchange between plants and their environment. In Arabidopsis thaliana, stomatal development is preceded by asymmetric cell divisions, and stomatal distribution follows the one-cell spacing rule, reflecting the coordination of cell fate specification. Stomatal development and patterning are regulated by both genetic and environmental signals. Here, we report that Arabidopsis MITOGEN-ACTIVATED PROTEIN KINASE3 (MPK3) and MPK6, two environmentally responsive mitogen-activated protein kinases (MAPKs), and their upstream MAPK kinases, MKK4 and MKK5, are key regulators of stomatal development and patterning. Loss of function of MKK4/MKK5 or MPK3/MPK6 disrupts the coordinated cell fate specification of stomata versus pavement cells, resulting in the formation of clustered stomata. Conversely, activation of MKK4/MKK5-MPK3/MPK6 causes the suppression of asymmetric cell divisions and stomatal cell fate specification, resulting in a lack of stomatal differentiation. We further establish that the MKK4/MKK5-MPK3/MPK6 module is downstream of YODA, a MAPKKK. The establishment of a complete MAPK signaling cascade as a key regulator of stomatal development and patterning advances our understanding of the regulatory mechanisms of intercellular signaling events that coordinate cell fate specification during stomatal development.

What Causes Opposing Actions of Brassinosteroids on Stomatal Development?

“…When CA-YDA, a constitutively active YDA variant that probably acts through activation of the MAPK pathway, which phosphorylates and inactivates SPCH (Wang et al, 2007;Lampard et al, 2008), was expressed in bri1-116, bsu-q, or bin2-1 mutants, stomatal development was blocked (Kim et al, 2012). Consistent with these data, the bsu-q spch-3 mutant exhibited a phenotype similar to that of spch-3, that is, lacked stomata (Kim et al, 2012).…”

“…Moreover, the overexpression of a mutant version of MKK4 in Thr-234, by replacing Thr with Ala to prevent phosphorylation, conferred a cotyledon epidermis composed almost entirely of stomata (Khan et al, 2013). This phenotype mirrors those in which the activities of either MKK4 and MKK5 or MPK3 and MPK6 are lost (Wang et al, 2007), which suggests that the MKK4 mutant version favorably competes with the endogenous one in binding MPK3 and MPK6, interfering with the activation of MPK3 and MPK6 (Khan et al, 2013). This would prevent SPCH repression, triggering stomatal development.…”

“…These genes encode peptide ligands (epidermal patterning factor, or EPF), and receptor proteins (TOO MANY MOUTHS [TMM] and the ERECTA family [ERf] of receptor-like kinases [ER, ERL1, and ERL2]; Nadeau and Sack, 2002;Shpak et al, 2005;Torii, 2012). Genes functioning downstream of these receptors include those encoding the mitogen-activated protein kinase (MAPK) module, which consists of the MAPK kinase kinase YODA (YDA); the MAPK kinases MKK4, MKK5, MKK7, and MKK9; and the MAPKs MPK3 and MPK6 (Bergmann et al, 2004;Wang et al, 2007;Lampard et al, 2009). MPK3 and MPK6 phosphorylate and inactivate the basic helix-loophelix transcriptional factor SPEECHLESS (SPCH), blocking the initiation of stomatal development (MacAlister et al, 2007;Pillitteri et al, 2007;Lampard et al, 2008).…”

A Tale of Two Systems: Peptide Ligand-Receptor Pairs in Plant Development

“…During stomatal development, EPF peptide signals received by ERECTA family are also likely to be mediated via a MAPK pathway, containing the MAPKKK YODA (YDA), the MAPKKs MKK4/5, and the MAPKs MPK3/6 for proper stomatal distribution (Bergmann et al 2004;Wang et al 2007). Constitutive activation of these MAPK cascade components leads to an epidermis devoid of stomata, and their loss of function overwhelmingly produces stomatal clusters, indicating that this MAPK module negatively regulates stomatal development via inhibition of the activity of the bHLH transcription factor specifying stomatal differentiation (Bergmann et al 2004;Wang et al 2007).…”

“…Constitutive activation of these MAPK cascade components leads to an epidermis devoid of stomata, and their loss of function overwhelmingly produces stomatal clusters, indicating that this MAPK module negatively regulates stomatal development via inhibition of the activity of the bHLH transcription factor specifying stomatal differentiation (Bergmann et al 2004;Wang et al 2007). In vitro phosphorylation of SPCH and the important role of phosphorylation-dependent SPCH degradation by MPK3/6 have indeed been demonstrated (Lampard et al 2008).…”