A central question is how specificity in cellular responses to the eukaryotic second messenger Ca²⁺ is achieved. Plant guard cells, that form stomatal pores for gas exchange, provide a powerful system for in depth investigation of Ca²⁺-signaling specificity in plants. In intact guard cells, abscisic acid (ABA) enhances (primes) the Ca²⁺-sensitivity of downstream signaling events that result in activation of S-type anion channels during stomatal closure, providing a specificity mechanism in Ca²⁺-signaling. However, the underlying genetic and biochemical mechanisms remain unknown. Here we show impairment of ABA signal transduction in stomata of calcium-dependent protein kinase quadruple mutant plants. Interestingly, protein phosphatase 2Cs prevent non-specific Ca²⁺-signaling. Moreover, we demonstrate an unexpected interdependence of the Ca²⁺-dependent and Ca²⁺-independent ABA-signaling branches and the in planta requirement of simultaneous phosphorylation at two key phosphorylation sites in SLAC1. We identify novel mechanisms ensuring specificity and robustness within stomatal Ca²⁺-signaling on a cellular, genetic, and biochemical level.

SUMMARY: Plant elicitor peptides (Peps) are conserved regulators of defense responses and models for the study of damage‐associated molecular pattern‐induced immunity. Although present as multigene families in most species, the functional relevance of these multigene families remains largely undefined. While Arabidopsis Peps appear largely redundant in function, previous work examining Pep‐induced responses in maize (Zm) implied specificity of function. To better define the function of individual ZmPeps and their cognate receptors (ZmPEPRs), activities were examined by assessing changes in defense‐associated phytohormones, specialized metabolites and global gene expression patterns, in combination with heterologous expression assays and analyses of CRISPR/Cas9‐generated knockout plants. Beyond simply delineating individual ZmPep and ZmPEPR activities, these experiments led to a number of new insights into Pep signaling mechanisms. ZmPROPEP and other poaceous precursors were found to contain multiple active Peps, a phenomenon not previously observed for this family. In all, seven new ZmPeps were identified and the peptides were found to have specific activities defined by the relative magnitude of their response output rather than by uniqueness. A striking correlation was observed between individual ZmPep‐elicited changes in levels of jasmonic acid and ethylene and the magnitude of induced defense responses, indicating that ZmPeps may collectively regulate immune output through rheostat‐like tuning of phytohormone levels. Peptide structure–function studies and ligand–receptor modeling revealed structural features critical to the function of ZmPeps and led to the identification of ZmPep5a as a potential antagonist peptide able to competitively inhibit the activity of other ZmPeps, a regulatory mechanism not previously observed for this family.

Brandt B, Munemasa S, Wang C, Nguyen D, Yong T, Yang PG, Poretsky E, Belknap TF, Waadt R, Alemán F, Schroeder JI. 2015. Calcium specificity signaling mechanisms in abscisic acid signal transduction in Arabidopsis guard cells. eLife 4:e03599. doi: 10.7554/eLife.03599