Phospholipase-Mediated Stress Response in Fission Yeast

裂殖酵母中磷脂酶介导的应激反应

• 批准号: 7320943
• 负责人: STEVAN MARCUS
• 金额: $ 8.83万
• 依托单位: UNIVERSITY OF ALABAMA IN TUSCALOOSA
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-03-01 至 2009-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7320943
• 关键词: Schizosaccharomyces pombe; adenylate cyclase; biological signal transduction; cell cycle; cell growth regulation; cell osmotic pressure; cyclic AMP; enzyme mechanism; fungal genetics; fungal proteins; gene mutation; growth media; lysophospholipase; microorganism growth; protein localization; stress

DESCRIPTION (provided by applicant): Phospholipase B (PLB) enzymes comprise what is perhaps the most poorly understood class of phospholipases in terms of biological functions. Although several PLB isozymes have been identified in genetically tractable yeasts, their physiological functions are largely unknown. We have elucidated a role for a newly identified member of the fungal PLB family, Plbl, as an essential mediator of osmotic stress response in the fission yeast, Schizosaccharomyces pombe, plb 1delta mutants exhibit mitotic and cytokinesis defects under conditions of osmotic stress that bear resemblance to defects observed in proteasome-defective mutants of S. pombe. They are also able to mate on rich media, a phenotype characteristic of mutants defective in function for the cAMP pathway. These phenotypes are distinct from those caused by loss of function of previously characterized stress response pathways in S. pombe, suggesting that Plb1 regulates a novel pathway. Consistent with the phenotypes caused by loss of Plb1 function, a screen for multi-copy suppressors of the osmosensitive growth defect of plb1 mutant cells resulted in the isolation of genes encoding activators of adenylate cyclase, a 26S proteasome subunit (pra7), and several genes encoding novel proteins, several of which are conserved in mammals. We have determined that the cAMP pathway, like PIb1, is essential for survival of S. pombe cells in hyperosmotic media and that loss of function of the cAMP pathway is rescued by several multi-copy suppressors of plb1delta, including pra7. Our findings lead us to hypothesize that PIb1 functions, in part, to regulate cell cycle progression under conditions of osmotic stress through a novel cAMP and 26S proteasomemediated pathway. We will use the power of yeast genetics, in combination with molecular and biochemical approaches, to further characterize the Plb1-dependent stress response pathway(s) in S. pombe. Our Specific Aims are to: (1) define the enzymatic activities and subcellular localization of Plb1 and determine whether they are altered in response to osmotic stress, (2) establish functional relationships between Plb1 and previously characterized osmotic stress response pathways in fission yeast, (3) investigate roles for the cAMP pathway and 26S proteasome in mediating Plb1-dependent stress response functions, and (4) determine whether other multicopy suppressors of plb1? encode components of the PIb1-dependent stress response pathway. We anticipate that our proposed studies will provide insights into novel mechanisms of stress-induced signal transduction and cell cycle regulation that are likely to be conserved in higher organisms. The proposed project is relevant to several human diseases, including cardiovascular, inflammatory, neurodesenerative, and neoplastic diseases.

描述（由申请人提供）： 就生物学功能而言，磷脂酶 B (PLB) 可能是人们最了解的磷脂酶类别。尽管在遗传易处理的酵母中已鉴定出几种 PLB 同工酶，但它们的生理功能在很大程度上尚不清楚。我们阐明了真菌 PLB 家族新发现的成员 Plbl 作为裂殖酵母裂殖酵母中渗透应激反应的重要介质的作用，plb 1delta 突变体在渗透应激条件下表现出类似的有丝分裂和胞质分裂缺陷在粟酒裂殖酵母蛋白酶体缺陷突变体中观察到的缺陷。它们还能够在丰富的培养基上交配，这是 cAMP 途径功能缺陷的突变体的表型特征。这些表型与先前在粟酒裂殖酵母中表征的应激反应途径功能丧失引起的表型不同，表明 Plb1 调节一种新的途径。与 Plb1 功能丧失引起的表型一致，对 plb1 突变细胞渗透敏感生长缺陷的多拷贝抑制因子的筛选导致编码腺苷酸环化酶激活剂、26S 蛋白酶体亚基 (pra7) 和几个基因的基因的分离编码新的蛋白质，其中一些在哺乳动物中是保守的。我们已经确定，cAMP 通路与 PIb1 一样，对于粟酒裂殖酵母细胞在高渗介质中的生存至关重要，并且 cAMP 通路功能的丧失可以通过 plb1delta 的多个多拷贝抑制因子（包括 pra7）来挽救。我们的研究结果使我们推测，PIb1 的功能部分是通过一种新的 cAMP 和 26S 蛋白酶体介导的途径在渗透压条件下调节细胞周期进程。我们将利用酵母遗传学的力量，结合分子和生化方法，进一步表征粟酒裂殖酵母中依赖于 Plb1 的应激反应途径。我们的具体目标是：(1) 定义 Plb1 的酶活性和亚细胞定位，并确定它们是否因渗透应激而发生改变，(2) 建立 Plb1 与裂殖酵母中先前表征的渗透应激反应途径之间的功能关系，( 3) 研究 cAMP 通路和 26S 蛋白酶体在介导 Plb1 依赖性应激反应功能中的作用，以及 (4) 确定是否还有其他 plb1 多拷贝抑制因子？编码 PIb1 依赖性应激反应途径的成分。我们预计我们提出的研究将为应激诱导的信号转导和细胞周期调节的新机制提供见解，这些机制可能在高等生物体中保守。该项目与多种人类疾病相关，包括心血管疾病、炎症疾病、神经退行性疾病和肿瘤疾病。