Differential Effects of the Peptides Stomagen, EPF1 and EPF2 on Activation of MAP Kinase MPK6 and the SPCH Protein Level

Abstract

The positioning and density of leaf stomata are regulated by three secretory peptides, EPIDERMAL PATTERNING FACTOR 1 (EPF1), EPF2 and stomagen. Several lines of published evidence have suggested a regulatory pathway as follows. EPF1 and EPF2 are perceived by receptor complexes consisting of a receptor-like protein, TOO MANY MOUTHS (TMM), and receptor kinases, ERECTA (ER), ERECTA-LIKE (ERL) 1 and ERL2. These receptors activate a mitogen-activated protein (MAP) kinase module. MAP kinases phosphorylate and destabilize the transcription factor SPEECHLESS (SPCH), resulting in a decrease in the number of stomatal lineage cells. Stomagen acts antagonistically to EPF1 and EPF2. However, there is no direct evidence that EPF1 and EPF2 activate or that stomagen inactivates the MAP kinase cascade, through which they might regulate the SPCH level. Experimental modulation of these peptides in Arabidopsis thaliana would change the number of stomatal lineage cells in developing leaves, which in turn would change the expression of SPCH, making the interpretation difficult. Here we reconstructed this signaling pathway in differentiated leaf cells of Nicotiana benthamiana to examine signaling without the confounding effect of cell type change. We show that EPF1 and EPF2 are able to activate the MAP kinase MPK6, and that both EPF1 and EPF2 are able to decrease the SPCH level, whereas stomagen is able to increase it. Our data also suggest that EPF1 can be recognized by TMM together with any ER family receptor kinase, whereas EPF2 can be recognized by TMM together with ERL1 or ERL2, but not by TMM together with ER.

Auxin represses stomatal development in dark-grown seedlings via Aux/IAA proteins

“…Interestingly, auxin treatment was shown to activate a MAPK cascade in Arabidopsis roots, and it has been suggested that this induction is independent of stress associated with this treatment (Mockaitis and Howell, 2000). Apart from the EPFL-ERf pathway (Jewaria et al, 2013;Lee et al, 2012), the YDA MAPK cascade also integrates signals from the BR signalling cascade (Khan et al, 2013;Kim et al, 2012) and the light signalling pathway (Kang et al, 2009). Thus, the YDA MAPK cascade is a central switch responding to intrinsic and environmental signals in the decision of whether or not to form a stoma.…”

Mitogen-activated protein kinase cascades in signaling plant growth and development

“…It is currently unclear whether SDD1 is involved in the processing of these ligands. Biochemical studies have demonstrated that EPF1 and EPF2 can activate MPK6, which results in the phosphorylation and destabilization of SPCH and a subsequent decrease in stomatal lineage cells [66]. However, strict genetic evidence to place them in the same linear pathway is still lacking.…”

Origins and Evolution of Stomatal Development

“…1H; Hara et al, 2009;Hunt and Gray, 2009;Hunt et al, 2010;Kondo et al, 2010;Sugano et al, 2010;Lee et al, 2012Lee et al, , 2015. Later in the stomatal lineage, EPF1 interacts with ERECTAs (primarily ERL1), again possibly overseen by TMM, to prevent GMC transitioning (Hara et al, 2007;Lee et al, 2012;Jewaria et al, 2013;Qi et al, 2017). This prevents neighboring cells from becoming stomata and promotes appropriate stomatal patterning and spacing.…”