Nitric Oxide and Autophagy Regulation in Arabidopsis

强大的稳态机制是细胞存活的关键。细胞必须保持稳定和最佳的细胞内条件，以进行熟练的酶和代谢过程。然而，一个高度动态和经常紧张的环境不可避免地导致细胞干扰。胞质游离钙和活性氧的增加是许多环境应激引起的细胞内变化的例子。细胞已经进化出了利用这些变化作为第二信使的机制。钙和氧化还原传感器蛋白感知这些压力信号，并传递信息，不仅激活细胞和生理适应性反应，而且还平衡机制，以恢复细胞内稳态。拟南芥CML24蛋白不仅是钙调素相关的钙离子传感器，而且可能是氧化还原传感器。CML24经历钙依赖性构象变化，并且还具有响应于氧化而形成二硫键的潜力。具有突变体CML 24的植物具有改变的一氧化氮积累和自噬调节。因此，一氧化氮和自噬调节在植物中可能是功能性相关的，并且这种联系对于细胞内稳态可能是至关重要的。钙和氧化还原在CML 24构象、稳定性、蛋白质相互作用和生理功能中的作用将通过生物化学、生物物理学和分子遗传学方法来确定。 将通过部分确定CML 24是否在NOA1-、硝酸还原酶-和/或α-依赖性一氧化氮合成途径中起作用来研究CML 24在一氧化氮积累中的作用。将监测自噬以揭示CML 24及其钙结合能力和二硫键形成是否对调控至关重要。一氧化氮积累和自噬突变体，除了cml24突变体，将其特征在于确定潜在的一氧化氮，氧化还原和自噬regulation.Broader ImpactsThis研究之间的联系将解决有关细胞应激感知和反应的基本和重要的问题，并可能带来变革性的洞察钙，氧化还原，一氧化氮和自噬在细胞内稳态的作用。这些过程对于植物的存活和生产力至关重要。从这些研究中获得的知识应有助于旨在提高农业效率和保护压力下的自然环境的做法。通过本科生、研究生和博士后的参与，本研究将加强研究和培训的基础设施。将鼓励获得该补助金支持的学生参加会议和教学机会。培训妇女和代表性不足群体的学生将是一个高度优先事项。将继续开展实验室图尔斯参观、科学博览会评审和NSF Advance活动等外联活动。研究成果将通过出版物、会议介绍和网站广泛传播。突变体和转基因株系将免费向社区开放。这项工作还涉及赖斯大学，美国各地和国际上的合作。

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 ImpactsThis 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.