Arabidopsis Actin Associated Protein SCRP1 Functions In Regulating Stomatal Movement

Actin cytoskeleton is highly-conserved and ubiquitous in eukaryotic cells. It forms filamentous network and constantly undergoes dynamic changes including assembly and disassembly processes. Actin dynamics functions in numerous cellular processes such as cell division, cell morphogenesis, cell tip growth and cell movement. Recently study reveals actin dynamics also plays roles in plant responses to abiotic or biotic stimuli. Actin dynamics is regulated by an array of actin binding proteins and actin associated proteins.Stomatal movement is important for plant to respond to exogenous stimuli in the environment. Actin dynamics plays vital roles in stomatal movement regulation. A great group of studies shows stomatal aperture is tightly correlated with actin filaments arrays, orientations, density and the extent of actin filaments bundling.In this study, a novel Arabidopsis gene SCRP1(stomatal closing regulation protein 1) was identified as a regulator functions in stomatal closing process. This study took forward genetic tool to perform a mutant screen using the leaves detached from T-DNA insertion lines ordered from ABRC to identify the mutant hypersensitive to water loss treatment. We identified a T-DNA mutant scrp1 which has faster water loss ratio from the screen. The T-DNA of the mutant is inserted into Arabidopsis gene SCRP1 which encodes a Ser/Thr protein kinase. In this study, we identified and characterized an Arabidopsis gene SCRP1, which responds to ABA treatment and participates in ABA-induced stomatal closure. Although SCRP1 does not bind to actin filaments directly or affect actin dynamics in vitro, it co-localizes with actin filaments and stabilizes actin filaments. Further investigation reveals that SCRP1 physically interacts with and phosphorylates Actin Depolymerizing Factor 4(ADF4) and inhibits its actin filament depolymerizing activity. During ABA-induced stomatal closure, deletion of SCRP1 in Arabidopsis impairs the repolymerization of actin filaments and renders stomatal closure more insensitive to ABA, whereas deletion of ADF4 impairs the disassembly of actin filaments and results in stomatal closure that is more sensitive to ABA and overexpression of ADF4 resulted in stomatal closure insensitive to ABA. Loss of function of ADF4 partially suppresses the stomatal closure phenotype in scrp1. Thus, our results suggest that SCRP1 regulates actin filament stability and stomatal closure mainly through phosphorylation of ADF4. The innovations of this thesis were shown as follows:1. Previous study demonstrated that Actin Depolymerizing Factor(ADF) is mainly responsible for actin disassembly and plays vital roles in numerous plant physiological processes including pollen tube elongation, hypocotyl growth, and guard cell movement. The activity of ADF is regulated by phosphorylation. This study finds that protein kinase SCRP1 phospohrylates ADF in Arabidopsis. These findings will make a great contribution to further study the physiological roles of ADF in plant.2. Recent evidence supports the involvement of actin cytoskeleton dynamics in regulation of stomatal closing. Actin cytoskeleton dynamics is regulated by numerous actin binding proteins and actin associated factors. However little is known about the actin associated protein kinase involved in stomatal closing modulation through regulating actin binding protein’s activity to actin. This study first indicates a novel actin associated protein kinase SCRP1 as an actin dynamics modulating protein regulating stomatal movement through phosphorylating ADF and inhibiting ADF actin severing activity. This contributes a lot to the study of actin dynamics involvement in stomatal movement regulation.