Robust homeostastic mechanisms are key to cellular survival. Cells must maintain stable and optimal intracellular conditions for proficient enzymatic and metabolic processes. However, a highly dynamic and often stressful environment inevitably leads to cellular disturbances. Increases in cytosolic free calcium and reactive oxygen species are examples of intracellular changes that result from many environmental stresses. Cells have evolved mechanisms to use these changes as second messengers. Calcium and redox sensor proteins perceive these signs of stress and convey information that activates not only cellular and physiological adaptive responses but also counterbalancing mechanisms to regain cellular homeostasis. The Arabidopsis CML24 protein may be not only a calmodulin-related calcium sensor but also a redox sensor. CML24 undergoes calcium-dependent conformational changes and also has the potential to form a disulfide bond in response to oxidation. Plants with mutant CML24 have altered nitric oxide accumulation and autophagy regulation. Thus, nitric oxide and autophagy regulation may be functionally linked in plants and such links may be critical for cellular homeostasis. The roles of calcium and redox in CML24 conformation, stability, protein interaction, and physiological function will be determined through biochemical, biophysical, and molecular genetic approaches. CML24's role in nitric oxide accumulation will be investigated through determining, in part, whether CML24 functions in an NOA1-, nitrate reductase-, and/or mitochondria-dependent nitric oxide synthesis pathway. Autophagy will be monitored to reveal whether CML24, and its calcium-binding ability and disulfide bond formation, are critical for regulation. Nitric oxide accumulation and autophagy mutants, in addition to cml24 mutants, will be characterized to determine potential linkages among nitric oxide, redox, and autophagy regulation.
Broader Impacts This research will address fundamental and important questions about cellular stress perception and response and may bring transformative insight into the roles of calcium, redox, nitric oxide, and autophagy in cellular homeostasis. Such processes are critical for plant survival and productivity. Knowledge derived from these studies should aid practices aimed at improving agricultural efficiency and preserving natural environments under stress. Through the involvement of undergraduates, graduate students, and postdocs, this research will enhance the infrastructure of research and training. Students supported by this grant will be encouraged to participate in conferences and teaching opportunities. Training of women and students of underrepresented groups will be a high priority. Outreach activities, such as lab tours, science fair judging, and NSF Advance activities, will continue. Research results will be disseminated widely through publication, conference presentations, and websites. Mutants and transgenic lines will be freely accessible to the community. This work also involves collaboration at Rice University, across the US, and internationally.
