Relationship of Antibiotic Production and Development in Streptomyces coelicolor

描述（由申请人提供）：本提案的主要目的是阐明一些复杂的，相互关联的抗生素生产过程和模式细菌颜色链霉菌形态发育之间的分子联系。链霉菌是一种非常有价值的生物活性分子来源，包括临床上使用的大多数抗生素。这些菌丝细菌通常合成抗生素和其他次生代谢物，同时发生更广泛的发育事件，导致菌落表面产生气生菌丝，其丝状细胞最终形成孢子。本研究的目的是关注两种特殊类型的蛋白质，它们影响着葡萄球菌抗生素的生产和发展。首先，磷酸蚁甲酰基转移酶是催化许多抗生素生物合成酶活性所需的翻译后修饰的酶。cocolicolor中SCO6673 PPTase基因的突变会导致产生钙依赖性抗生素的能力丧失，但反过来也会导致潜在抗癌分子十一基芥子红素的过量产生。在这个突变体中，产孢也延迟了。为了了解这些不同表型效应的基础，实时PCR将用于评估PPTase突变体中已知抗生素调控和发育基因的表达与野生型相比的任何差异。这些数据将为随后的靶向突变策略提供信息，以设计S. coelicolor以最大限度地产生十一烷基芥子红素。第二个目标是了解应激反应中ECF sigma因子的活动。U可以阻断聚酮类抗生素的产生和气生菌丝的形成。微阵列分析和蛋白质组学研究的结合比较了野生型和rsuA突变体的高，不调节？活动将建立完整的规则？依赖u的基因以及间接受？你的活动。在rsuA突变体中观察到的细胞外蛋白酶和戊糖磷酸途径酶活性的升高与其抗生素生产和发育缺陷的相关性将通过相关基因的突变或过表达以及对所产生菌株的分析来测试。突变分析也将用于测试可能的膜蛋白SCO4110在调节？你的活动。更好地了解次级代谢和发育是如何相互影响的，可能最终为链菌有效的药物生产提供可推广的策略。

DESCRIPTION (provided by applicant): The broad objective of this proposal is to elucidate some of the molecular connections between the complex, interrelated processes of antibiotic production and morphological development in the model bacterium Streptomyces coelicolor. The streptomycetes are an incredibly valuable source of bioactive molecules, including the majority of antibiotics clinically used. These mycelial bacteria typically synthesize antibiotics and other secondary metabolites concurrently with broader developmental events that result in the generation of an aerial mycelium on the colony surface whose filamentous cells ultimately sporulate. The aims of this study concern two particular types of proteins that influence both antibiotic production and development in S. coelicolor. First, the phosphopantetheinyl transferase are enzymes that catalyze a posttranslational modification required for the activity of many antibiotic biosynthetic enzymes. Mutation of the SCO6673 PPTase gene in S. coelicolor results in loss of the ability to produce calcium dependent-antibiotic but also conversely hyperproduction of the potential anti-cancer molecule undecylprodigiosin. Sporulation is also delayed in this mutant. In order to understand the basis for these diverse phenotypic effects, real-time PCR will be used to assess any differences in expression of known antibiotic regulatory and developmental genes in the PPTase mutant as compared to the wild type. This data will inform a subsequent targeted mutational strategy to engineer S. coelicolor for maximum undecylprodigiosin production. The second aim focuses on understanding how activity of a likely stress response ECF sigma factor, ?U, can block both polyketide antibiotic production and aerial mycelium formation in S. coelicolor. A combination of microarray analysis and proteomic studies comparing wild type S. coelicolor to a rsuA mutant with high, unregulated ?U activity will establish the complete regulon of ?U-dependent genes as well as those indirectly affected by ?U activity. The relevance of the observed elevation of both extracellular protease and pentose phosphate pathway enzyme activity in the rsuA mutant to its antibiotic production and developmental defects will be tested by mutation or overexpression of the relevant genes and analysis of the resulting strains. Mutational analysis will also be used to test the involvement of the likely membrane protein SCO4110 in the regulation of ?U activity. A better understanding of how secondary metabolism and development are mutually influenced may ultimately suggest generalizable strategies for effective pharmaceutical production in the streptomycetes. PUBLIC HEALTH RELEVANCE: The goal of the proposed research is to understand how Streptomyces bacteria make useful compounds, such as antibiotics, as part of their overall life cycle. The better we understand how these beneficial bacteria grow, the more likely it is that we can engineer that growth to produce life-saving drugs for fighting infections and cancers.