Char of Phospholipases C & the Twin Arginine Secretory System of B. pseudomallei

介绍细胞内铁浓度的动态控制是至关重要的所有生物系统。 这个问题的一个方面是生物学上有用的铁（即Fe2-）是极其有限的或它是高度不溶性的 （即Fe 3）。因此，生物实体已经进化出有效的机制来从植物中获取这种营养素 不溶性形式，其通常是大量的。另一方面，进一步收购铁上面 生物学上有用的浓度可能对细胞产生可怕的后果。过量的游离铁会催化 产生高活性氧和氮中间体，这将破坏所有已知的生物 大分子这种冲突，在一个主要的方式是处理在不同的致病微生物阵列， 一种典型的依赖于Fe 2+的阻遏物，称为Fur（铁吸收调节剂），在控制铁中起关键作用 体内平衡虽然很明显B类选择性病原体类鼻疽伯克霍尔德氏菌和鼻疽伯克霍尔德氏菌表达 与来自密切相关的生物体（例如铜绿假单胞菌）的毛皮高度同源的蛋白质， 关于铁稳态在8的毒性中的作用知之甚少。假鼻疽和鼻疽。更 重要的是，对于铁稳态在这些细胞的细胞内生活方式中的作用， 病原体我们建议通过使用强有力的分子和遗传技术来补救这种情况。 我们采用的方法来阐明毛皮在相关的，但 独特的机会致病菌铜绿假单胞菌。我们将确定6的Fur调控基因。假鼻疽& 编码调节因子（例如σ因子、小调节RNA）的鼻疽。然后我们将评估 这些选定的调节因子在这些因子进入、存活、生长或运输的能力中的作用 模型真核细胞（如巨噬细胞），适用于已知的发病机制 这些潜在的生物恐怖分子我们鉴定的基因产物最终可能会用于 开发针对类鼻疽和腺体的治疗和预防剂。 项目互动除了与其他项目成员（例如Vazquez博士-）的明显互动之外 托雷斯，Holmes，& Voskuil），其在本申请的主体中以及在附图中更详细地描述。 介绍项目II. C，我们将特别与罗宾逊博士（杨百翰大学选择代理档案）和博士。 Schweizer（微生物遗传学-CSU）在细菌学和遗传学方面的B。假鼻疽和鼻疽。 我们也将特别感兴趣的是非常积极的互动与博士Slayden（基因组学/蛋白质组学 核心），这将是至关重要的发展微阵列和评估潜在的靶基因， 可能用于实验性预防和治疗剂的产品。此外，我们将沿着这些路线 也与Brennan博士（PDM核心）密切互动，以达到同样的目的。

Introduction The dynamic control of intracellular iron concentrations is paramount to all biological systems. One aspect of this issue is that biologically useful iron (i.e. Fe2") is extremely limiting or it is highly insoluble (i.e.Fe3). Accordingly, biological entities have evolved efficient mechanisms to acquire this nutrient from the insoluble form, which is generally in plentiful quantities. On the other hand, further acquisition of iron above biologically useful concentrations can have dire consequences for a cell. Excess free iron will catalyze the generation of highly reactive oxygen and nitrogen intermediates that will damage all known biological macro-molecules. This conflict, in a major way is dealt with in a diverse array of pathogenic microbes, by a classic Fe2+-dependent represser called Fur (ferric uptake regulator) that plays the key role in controlling iron homeostasis. While it is clear that the Class B Select Agents Burkholderia pseudomallei & mallei express a protein that is highly homologous to Fur from closely related organisms (e.g. Pseudomonas aeruginosa) very little is known about the role of iron homeostasis in the virulence of 8. pseudomallei & mallei. More importantly, nothing is known about the role of iron homeostasis in the intracellular life style of these pathogens. We propose to remedy this situation through the use of powerful molecular and genetic approaches that we employed to elucidate the extensive and crucial role that Fur plays in the related, but distinct, opportunistic pathogen P. aeruginosa. We will identify the Fur regulated genes of 6. pseudomallei & mallei that encode regulatory factors (e.g. sigma factors, small regulatory RNAs). We will then evaluate the role of these selected regulatory factors in the ability of these agents to enter, survive in, grow in or traffick in model eukaryotic cells (e.g. macrophage), which are appropriate to the known aspects of the pathogenesis of these potential bioterrorist agents. The gene products that we identify may ultimately be used in developing therapeutic and prophylactic agents against Malleipdosis and Glanders. Project interactions In addition to the obvious interactions with other project members (e.g. Drs. Vazquez- Torres, Holmes, & Voskuil) that are described in more detail in the body of this application and in the Introduction to Project II.C, we will especially interact with Dr. Robison (BYU Select Agent Archive) and Dr. Schweizer (Microbial genetics - CSU) in terms of the bacteriology and genetics of B. pseudomallei & mallei. We will also be especially interested in very active interactions with Dr. Slayden (Genomics/Proteomics Core) that will be vital to development of microarrays and for the evaluation of potential target genes and products for possible use in experimental prophylactic and therapeutic agents. Also, along these lines we will also closely interact with Dr. Brennan (PDM Core) for the same purpose.