Dr. Melissa Bredow Postdoctoral Fellow, Monaghan Lab, Queen's University Exploring site-specific modification as a tool to direct pathway specific activity of the multifunctional calcium-dependent protein kinase CPK28
Calcium (Ca2+) is a ubiquitous secondary messenger involved in most adaptive and developmental signaling programs in plants. Spatially and temporally defined influxes of intracellular Ca2+, known as Ca2+ signatures, are generated in response to environmental cues and are “decoded” by intracellular proteins. Ca2+-dependent protein kinases (CDPKs) are a unique family of Ca2+ sensors that can both perceive Ca2+ and propagate intracellular signals. Many CDPKs serve roles in several pathways and therefore must be capable of decoding distinct Ca2+ signatures. How stimuli specific Ca2+ signatures are distinguished has remained an outstanding and compelling question. In our work, we explore site-specific phosphorylation as a regulator of Ca2+ sensitivity on Arabidopsis CPK28. We show that phosphorylation at an individual residue (Ser318) dictates the level of Ca2+ required for kinase activation and directs pathway-specific activity of CPK28 in vivo. CPK28 is a negative regulator of immune signaling and also functions in the vegetative-to-reproductive stage transition. Generating a CPK28 allele that could not be phosphorylated at Ser318 resulted in enhanced immunity to bacterial infection without consequences to reproductive growth observed in cpk28 loss-of-function plants. Biochemical analysis indicated that Ser318 phosphorylation promoted a change in protein conformation that “primed” CPK28 for rapid Ca2+ activation, necessary for immune function. In contrast, Ser318 phosphorylation was not required for reproductive growth indicating pathway specific requirements for rapid Ca2+ activation. Further, we identify additional CPK28 phospho-mutant alleles that direct pathway specific activity, enhancing bacterial immunity and promoting plant growth. Ongoing work is aimed at understanding the evolution of multifunctionality in CDPKs and suggests that CPK28 belongs to an ancient core immune pathway. Together, our work points towards a conserved mechanism of Ca2+ priming in CPK28 orthologs and highlights the use of the site-specific modifications to generate stress-resilient plants without fitness costs.