Forespore Engulfment During B. subtilis Sporulation

描述（由申请人提供）：芽孢杆菌属和梭状芽孢杆菌属的细菌产生异常持久和长寿的孢子，这些孢子是炭疽和肉毒杆菌中毒的传染因子，并在另一个细胞的细胞质中组装。这种独特的细胞结构是由类似吞噬作用的吞噬过程产生的，在这个过程中，较大的母细胞的膜围绕较小的前孢子迁移，直到它完全包裹在母细胞的细胞质中。吞噬为细菌细胞的动态能力提供了一个戏剧性的例子，但其机制尚不清楚。在此之前，仅有的吞噬突变体阻断了室间隔变薄，在此期间，室间隔内的肽聚糖变薄，为膜迁移做准备。我们已经开发了新的工具来研究吞噬，并确定了在膜迁移中有缺陷的突变体，并在吞噬的最后一步，膜融合。膜融合缺陷突变体影响一种在许多物种中高度保守和必需的蛋白质。该蛋白定位于分裂位点，参与染色体分离的最后阶段，这表明它也可能参与细胞分裂完成时的膜融合，这一过程鲜为人知。在间隔变薄过程中，水解肽聚糖需要产孢特异性酶，我们将测试营养自溶素是否可以部分替代产孢特异性酶。自溶酶存在于所有细菌中（据预测枯草芽孢杆菌基因组编码超过30种这样的酶），并且被认为允许肽聚糖重塑以促进细胞的延伸和分裂。然而，这些酶具有潜在的致命性，因为除非它们在空间和时间上受到严格的调控，否则它们的活性会导致细胞裂解。事实上，许多商业抗生素的致命性需要自溶素。吞噬为理解细菌如何控制这些潜在的致命酶提供了一个理想的系统，这些酶是新型抗生素的有吸引力的靶标。我们将采取细胞生物学、遗传学和生物化学相结合的方法，研究肽聚糖水解的空间调控，细菌细胞膜融合的机制，以及细菌如何在细胞内移动和定位大分子

DESCRIPTION (provided by applicant): Bacteria from the genera Bacillus and Clostridium produce unusually durable and long-lived spores that are the infectious agent of Anthrax and Botulism, and which are assembled in the cytoplasm of another cell. This unique cell within a cell structure is produced by the phagocytosis-like process of engulfment, during which the membrane of the larger mother cell migrates around the smaller forespore, until it is completely enclosed within the mother cell cytoplasm. Engulfment provides a dramatic example of the dynamic capabilities of the bacterial cell, but its mechanism remains unclear. Previously, the only engulfment mutants blocked septal thinning, during which peptidoglycan within the septum is thinned in preparation for membrane migration. We have developed new tools for the study of engulfment, and identified mutants defective in membrane migration, and in the final step of engulfment, membrane fusion. The membrane fusion defective mutants affect a protein that is both highly conserved and essential in many species. This protein localizes to site of division and is involved in the final stages of chromosome segregation, suggesting that it may also be involved in membrane fusion at the completion of cell division, a process about which little is known. Sporulation-specific enzymes are required to hydrolyze peptidoglycan during septal thinning, and we will test if vegetative autolysins can partially substitute for the sporulation specific enzymes. Autolysins are found in all bacteria (the Bacillus subtilis genome is predicted encode more than 30such enzymes), and are thought to allow peptidoglycan remodeling for cell elongation and division. However, these enzymes are potentially lethal, since unless they are tightly regulated both spatially and temporally, their activity can result in cell lysis. Indeed, the lethality of many commercial antibiotics requires autolysins. Engulfment provides an ideal system for understanding how bacteria control these potentially lethal enzymes, which are attractive targets for novel antibiotics. We will take a combined cell biological, genetic and biochemical approach to study the spatial regulation of peptidoglycan hydrolysis, the mechanism of membrane fusion in bacterial cells, as well as to understand how bacteria move and localize macromolecules within their cells