Centromere structure and isochromosome formation in Candida albicans

描述（由申请人提供）：白色念珠菌是最普遍的人类真菌病原体，能够引起侵袭性念珠菌病，这是一种死亡率高、医疗费用高的严重疾病。氟康唑是治疗念珠菌感染最常用的口服抗真菌药物，氟康唑获得性耐药（FluR）是一个重要问题。非整倍体是由涉及着丝粒的非分离事件引起的，与FluR密切相关。与高等真核生物一样，白色念珠菌着丝粒具有独特的区域性，但与单个微管相连，缺乏像酿酒酵母点着丝粒一样的中心周围异染色质。对白色念珠菌着丝粒及其相关着丝点蛋白组分的研究将有助于更好地了解CEN的结构和功能。这一知识可以用于直接抗真菌药物的鉴定，以及通过改变CEN功能来改变非整倍体频率和FluR获得的伴随药物的开发。该项目将表征白色念珠菌着丝点的结构，重点是DASH蛋白复合物的成员。酿酒酵母和裂糖酵母的DASH复合体成员的预测同源物将着丝粒附着在微管上。重要的是，DASH复合体蛋白在许多致病真菌中是保守的，在后生动物中缺乏同源物。在酿酒葡萄球菌中，DASH复合体成员对生长至关重要，可能是有希望的药物靶点。本研究旨在探讨DASH复合体成员在白色念珠菌中的定位、DASH复合体成员与其他着丝点蛋白之间的相互作用，以及DASH复合体成员对白色念珠菌生长的重要性。此外，本项目将验证在某些白色念珠菌CEN区域发现的反向重复DNA序列有助于片段性非整倍体的发展，从而获得耐药性的假设。总之，拟议的实验将表征DASH复合物蛋白作为潜在的抗真菌药物靶点，并深入了解片段性非整倍体形成的机制，这一过程与氟康唑耐药性的发展相关。公共卫生相关性：白色念珠菌是最常见的分离的人类真菌病原体，对治疗过程中使用的抗真菌药物的耐药性是一个重大问题。本研究将研究几种与白色念珠菌着丝粒相关的蛋白质，以表征它们作为抗真菌药物靶点的潜力。此外，该项目将深入了解导致对常用抗真菌药物产生耐药性的特定染色体重排的机制。

DESCRIPTION (provided by applicant): Candida albicans is the most prevalent human fungal pathogen and is capable of causing invasive candidiasis, a severe disease with high mortality rates and medical costs. Fluconazole is the most frequently prescribed oral antifungal for treating Candida infections and acquired resistance to fluconazole (FluR) is a significant problem. Aneuploidies, which arise from non-disjunction events that involve centromeres, are strongly correlated with FluR. C. albicans centromeres are unique in being regional, like those of higher eukaryotes, yet attaching to a single microtubule and lacking pericentric heterochromatin, like Saccharomyces cerevisiae point centromeres. Characterization of C. albicans centromeres and the kinetochore protein components associated with them will provide a better understanding of CEN structure and function. This knowledge can be exploited for the identification of direct anti-fungal drugs as well as for the development of companion drugs that alter the frequency of aneuploidies and the acquisition of FluR,by altering CEN function. This project will characterize the structure of the C. albicans kinetochore, focusing on members of the DASH protein complex. Predicted homologs of DASH complex members in S. cerevisiae and Schizosaccharomyces pombe attach centromeres to microtubules. Importantly, DASH complex proteins are conserved in many pathogenic fungi and lack homologs in metazoans. In S. cerevisiae, DASH complex members are essential for growth and may be promising drug targets. This proposal addresses the localization of DASH complex members in C. albicans, the interactions between DASH complex members and other kinetochore proteins, and the essentiality of DASH complex members for C. albicans growth. In addition, this project will test the hypothesis that the inverted repeat DNA sequences found in some C. albicans CEN regions contribute to the development of segmental aneuploidies and thus to the acquisition of drug resistance. Together, the proposed experiments will characterize the DASH complex proteins as potential antifungal drug targets and gain insight into the mechanism of segmental aneuploidy formation, a process that is correlated with the development of resistance to fluconazole. PUBLIC HEALTH RELEVANCE: Resistance of Candida albicans, the most commonly isolated human fungal pathogen, to anti- fungal drugs used during treatment is a significant problem. This proposal will examine several proteins associated with the C. albicans centromere to characterize their potential as anti-fungal drug targets. Also, this project will gain insight into the mechanism of how specific chromosomal rearrangments that result in resistance to a commonly used anti-fungal drug arise.