Functional Analysis of the High-Light-Inducible Proteins (HLIP) in Synechocystis PCC 6803

智慧的光是光合作用所必需的，光合作用是地球上几乎所有生物的食物的最终能量来源。然而，当光合作用细胞吸收的光能超过光合作用能力时，细胞内会产生活性氧物种，对蛋白质、脂质和光合色素造成氧化损伤。这些有害影响被其他环境压力(如低温、干旱)放大。在次优生长条件下，植物或藻类无法有效利用光合作用产生的还原能量。这降低了过量的光吸收导致光损伤的光强度。因此，在所有其他胁迫条件下，光合作用生物也不可避免地面临光胁迫。然而，尽管光胁迫与作物生产力有关，但植物或光合作用微生物对光胁迫反应的分子机制尚未阐明。该项目的目标是确定光合作用生物保护自己免受高强度光伤害的分子机制。本研究重点分析了模式生物聚球藻PCC 6803中四个高光诱导蛋白家族(HLIP)的功能。HLIP在其初级序列上类似于高等植物的捕光叶绿素a/b结合蛋白。它们已被证明是蓝藻在强光下生存所必需的。它也可能在高等植物中发挥关键作用。为了研究HLIPs及其相关蛋白在强光下细胞存活中的作用，将对PfsR(光合作用、铁稳态和胁迫反应调节)进行进一步的表征，以了解其在强光下的保护作用。在五个在强光下恢复活性的抑制子中的至少一个中的第二位点突变将被精确定位到四重hli缺失突变体。这些研究将揭示细胞在强光下存活的机制和HLIP的功能。此外，HLIP与光系统I三聚体相互作用的影响以及色素结合基序在HLIP中的作用将通过定点突变来确定。此外，类囊体膜上一种新的高光诱导类胡萝卜素结合蛋白复合体将被表征。总体而言，这项研究将提高对光合作用生物在强光条件下生存机制的理解，并最终提高为植物或藻类制定合理策略以提高抗逆性的能力。广泛影响植物生物学家普遍对该研究课题感兴趣。这项研究使学生和博士后研究员能够将遗传学、生物化学和生物物理学的方法整合到他们的教育和研究计划中，培养对农业和生物燃料生产至关重要的具有高度竞争力和创新性的研究计划。它将使受训人员为独立研究生涯做好准备。此外，它还将提高阿肯色州的光合作用研究力度。该项目将使国际学生联合会继续让代表人数不足的少数族裔学生参与研究，有助于他们继续接受高等教育。约35%的UALR本科生(超过4000名学生)来自代表性较低的群体，但由于实验室空间和活跃的教师研究项目的短缺，很少有人有机会进行研究培训。此外，该项目还将通过州或UALR资助的涉及少数族裔学生的项目，为本科生的培训提供额外资金。

Intellectual MeritLight is required for photosynthesis, which is the ultimate source of energy in food for almost all living things on earth. However, when the light energy absorbed by photosynthetic cells exceeds the photosynthetic capacities, reactive oxygen species are generated in the cells, causing oxidative damage to proteins, lipids, and photosynthetic pigments. These detrimental effects are amplified by other environmental stresses (e.g. low temperature, drought). Under suboptimal growth conditions plants or algae cannot efficiently use the reducing power generated by photosynthesis. This lowers the light intensity at which excess light absorption leads to photo-damage. Hence, photosynthetic organisms unavoidably face light stress as well during all other stress conditions. However, in spite of the importance of light stress in relation to crop productivity, the molecular mechanisms of light-stress responses have not yet been elucidated in plants or photosynthetic microbes. The goal of this project is to determine the molecular mechanisms by which photosynthetic organisms protect themselves from high intensity light. The research focuses on analysis of the functions of a family of four high light-inducible proteins (HLIP) in the model organism Synechocystis PCC 6803. HLIPs resemble the light-harvesting chlorophyll a/b-binding proteins of higher plants in their primary sequences. They have been proven to be essential for cyanobacteria to survive exposure to high light. It may also play a critical role in higher plants as well. To investigate the functions of HLIPs and associated-proteins in cell survival during exposure to high light, the regulator PfsR (photosynthesis, Fe homeostasis and stress-response regulator) will be further characterized in order to understand its roles in protection against high intensity light. The second-site mutation in at least one of the five suppressors that restore viability in high light to the quadruple hli deletion mutant will be pinpointed. These studies will reveal the mechanisms of cell survival and the functions of HLIPs in high light. In addition, the effects of HLIP and photosystem I trimer interaction and the role of the pigment-binding motifs in HLIPs will be defined by site-directed mutagenesis. Furthermore, a novel high light-inducible, carotenoid-binding protein complex in the thylakoid membrane will be characterized. Overall, the research will improve understanding of the mechanisms for survival of photosynthetic organisms under high light conditions and will eventually enhance the ability to develop rational strategies for engineering plants or algae for better stress tolerance.Broader ImpactThe research subject is of general interest to plant biologists. The research allows students and postdoctoral fellows to integrate approaches of genetics, biochemistry and biophysics in their educational and research programs, fostering highly competitive and novel research initiatives important to agriculture and biofuel production. It will prepare the trainees for independent research careers. In addition, it will improve the strength of photosynthesis research in Arkansas. The project will allow the PI to continue involving underrepresented minority students in research, contributing to their retention in higher education. About 35% UALR undergraduates (more than 4000 students) are from underrepresented groups, but few have opportunities for research training due to the shortage of laboratory space and active faculty research programs. Furthermore, this project will stimulate additional funding for the training of undergraduate students through State- or UALR-sponsored programs that involve minority students.