TWO-COMPONENT HISTIDINE KINASES OF CANDIDA ALBICANS

白色念珠菌是美国真菌感染的主要原因，念珠菌属（Candida spp.）在引起医院感染的病原体中居第四位。 该菌的毒力可能是多因子和组织特异性的，因为C。白色念珠菌能够感染许多不同的环境部位（例如，酸性阴道腔与中性pH的血液和组织）。 双组分组氨酸激酶（HK）是原核生物和低等真核生物的环境传感器、磷酸化蛋白，其活性导致与对胁迫的适应性反应相关的基因的转录激活。 在致病真菌的HK基因中，我们最近分离到一个C.白念珠菌杂合HK基因（CaHK 1）是迄今为止所描述的所有HK基因中结构和功能独特的。 一个cahk 1空菌株已被构建和比较表型在体外和体内的亲本和基因重建的菌株。 在体外，无效菌株的菌丝在pH7.5絮凝，表明菌丝中的细胞表面变化与caHK 1中的突变有关。与亲本和基因重建菌株相比，无效菌株在系统性鼠模型中也是无毒的，但在大鼠阴道粘膜模型中是有毒的. 虽然北方分析表明，CaHK 1在酸性和中性pH介质中表达，我们推测，Cahk 1 p的活性可能取决于其磷酸化状态。 该提案有三个具体目标。 1.将使用北方分析在体外测定环境因素（温度、pH值、渗透压摩尔浓度）对CaHK 1表达的影响;将测定Cahk 1 p的翻译后修饰。 2.我们将在口腔和阴道模型中评估CaHK 1菌株构建体的毒力，以扩展我们对Cahk 1 p在毒力中作用的认识。 3. Cahk 1 p与其他蛋白质，包括下游效应蛋白的相互作用有待鉴定。 总之，这些研究将部分确定Cahk 1 p磷酸化途径和功能的CaHK 1念珠菌病的发病机制。

Candida albicans is the leading cause of fungal infections in the US and, Candida spp. now rank fourth among all pathogens in causing nosocomial infections. Virulence in this organism is probably multi-factorial and tissue-specific, since C. albicans is capable of infecting a number of diverse environmental sites (for example, the acid vaginal canal vs. the neutral pH of blood and tissues). Two-component, histidine kinases (HK) are environmental sensor, phosphorelay proteins of prokaryotes and lower eukaryotes whose activity results in transcriptional activation of genes associated with an adaptive response to stress. Among the HK genes of pathogenic fungi, recently, we isolated a C. albicans hybrid HK (CaHK1), which is unique structurally and functionally among all HK genes thus far described. A cahk1 null strain has been constructed and compared phenotypically in vitro and in vivo to parental and gene-reconstituted strains. In vitro, hyphae of null strains flocculate at pH 7.5, indicating that cell surface changes in hyphae are associated with a mutation in caHK1 The null is also avirulent in a systemic murine model but virulent in a rat vaginal mucosal model when compared to parental and gene- reconstituted strains. While Northern analyses indicate that CaHK1 is expressed in both acid and neutral pH media, we hypothesize that activity of Cahk1p may depend upon its state of phosphorylation. There are 3 specific aims in this proposal. 1. The influence of environmental factors (temp, pH, osmolarity) on the expression of CaHK1 will be measured in vitro using Northern analyses; post-translational modifications of Cahk1p will be determined. 2. We will evaluate the virulence of CaHK1 strain constructs in oral and vaginal models in order to extend our knowledge on the role of Cahk1p in virulence. 3. Cahk1p interactions with other proteins including down-stream effector proteins are to be identified. In summary, these studies will partially define the Cahk1p phosphorelay pathway and function of CaHK1 in the pathogenesis of candidiasis.