Generation of Diverse Centrosomes, Cilia and Flagellae During Development

点击翻译按钮获取中文摘要

PROJECT SUMMARY Centrioles are required to template cilia and flagella and to organize centrosomes that nucleate cytoplasmic microtubules in interphase and dividing cells. Centrosomes lose their ability to organise cytoplasmic microtubules when they become basal bodies, which can template the formation of primary cilia for signalling or flagellae for propelling sperm. To understand these diverse roles, it necessary to study centrioles in the development of different cell types. Yet, their replication, structure and function has been largely studied in cultured cells. Here we take advantage of the genetic and developmental advantages of Drosophila to study centriole function in two cell types; cells destined to make neurosensory cilia in the leg imaginal disc and male germ line cells destined to generate sperm. Our focus is on the roles of a network of centriole proteins in which Ana3, counterpart of human ROTATIN and Rcd4, PPP1R35 in human cells, interact with each other and with a set of proteins required for various steps of the elongation and maturation of the centriole necessary for it to become a basal body. Specifically, we will: (i) apply molecular and structural biology approaches to determine how the Ana3 and Rcd4 proteins interact with each other and with other centriole proteins to specify different aspects of centriole assembly and function and how this is regulated by phosphorylation. We will test the functions of these interactions through directed mutagenesis. (ii) use genetic approaches to determine inter-dependencies of the networks that regulate centriole elongation in the conventional cell division cycles of cultured cells and the leg imaginal disc and in cells that develop neurosensory cilia. (iii) determine the differential roles of Rcd4 and Ana3 and their interacting network of proteins in the multiple steps leading to formation of the giant centrioles of spermatocytes and their sequential roles in generating primary cilia and subsequently sperm flagellae., Because centriole duplication pathways are highly conserved between Drosophila and man, our findings will shed light onto a wide range of human diseases including cancer, where centrosome defects indicate poor prognosis; heritable ciliopathies, which affect centrioles and cilia in a wide range of tissues; and in microcephaly, where defects in proteins such as ROTATIN, whose counterpart we study here, lead to abnormal brain development.