Analyzing allele specific gene expression and regulation in C. albicans.

背景：大多数真核生物是杂合的，由两个不同的等位基因组成。然而，对于不同的等位基因是如何差异表达和调节的，以及差异调节是否能赋予生物体适应性，人们知之甚少。这些研究将为研究等位基因特异性表达的程度、机制和功能提供重要信息。这些话题与理解癌症的机制高度相关。目的/假设：白色念珠菌的天然杂合性使其能够在哺乳动物宿主中进化和适应。然而，差异等位基因的表达程度及其调控机制尚未完全阐明。我建议确定差异等位基因表达的程度，并阐明在真菌二态条件下特异性转录因子的差异结合是否调节了这种差异等位基因的表达。这项工作有望为真核生物杂合性的程度和作用提供见解。具体目的：1)全面检查白色念珠菌中差异等位基因表达的程度。2)确定差异基因表达是否与Poll I与转录因子结合的差异有关。3)测定差异调控等位基因的功能。研究设计：拟以白色念珠菌为模型系统，研究差异等位基因的表达与调控。我将使用454种DNA测序方法对等位基因特异性基因表达进行全局分析。ChlPSeq。（染色质免疫沉淀和测序）将用于评估有多少是由于转录调节。最后，我将使用基因替代方法确定差异等位基因表达的功能。

DESCRIPTION (provided by applicant): Background: Most eukaryotes are heterozygous, comprised of two distinct alleles. However, little is known about how distinct alleles are differentially expressed and regulated and whether differential regulation can confer fitness to an organism. The proposed studies will provide important information as to the extent of allele specific expression, its mechanisms and function. These topics are highly relevant to understanding mechanisms of cancer. Objective/hypothesis: The natural heterozygosity of C. albicans has allowed it to evolve and adapt to thrive in its mammalian host. However, the degree of differential allelic expression and the mechanism of its regulation are yet to be fully elucidated. I propose to determine the extent of differential allelic expression, and to elucidate if differential binding of specific transcription factors under fungal dimorphic conditions regulates this differential allelic expression. This work is expected to provide insights into the extent and role of heterozygosity of eukaryotes in general. Specific Aims: 1) Globally examine the extent of differential allelic expression in Candida albicans. 2) Determine if differential gene expression is associated with differences in binding of Poll I and transcription factors. 3) Assay the function of differentially regulated alleles. Study Design: I plan to study differential allelic expression and regulation using C. albicans as a model system. I will use 454 DNA sequencing methods to globally analyze allele specific gene expression. ChlPSeq. (chromatin immunoprecipitation and sequencing) will be used to assess how much is due to transcriptional regulation. Finally, I will determine the function of differential allelic expression using gene replacement methods. Relevance: A hallmark of cancer is genome instability. This can be caused by hypomethylation and/or loss of heterozygosity. Moreover, it is common for specific chromosomes and specific chromosomal regions to be preferentially amplified. Thus, basic understanding of the diversity of allele specific expression and how it can be controlled is of high importance to cancer. Moreover, C. albicans is an opportunistic pathogen that affects many diseases, including cancer. Understanding mechanisms that control its growth and switch to dimorphic growth, which is involved in pathogenesis, is of significance for treatment of cancer patients.