Control of centriole length in C.elegans and Drosophila

The eukaryotic cell cycle consists of a tightly regulated series of events that ensure the correct duplication and segregation of the genetic material to each daughter cell. Errors in this choreography can lead to genetic instability and result in uncontrolled cell proliferation. The microtubule (MT) cytoskeleton plays a pivotal role in the correct segregation of DNA as it builds the mitotic spindle and organises the cleavage furrow. " Centrosome organisation and function during the cell cycle Centrosomes are found at the two poles of the mitotic spindle and serve as microtubule organising centres. Centrosomes consist of a centriole pair surrounded by the pericentriolar material. Centriole duplication is tightly regulated to ensure they duplicate only once per cell cycle. As cells enter mitosis, the centrioles recruit PCM (maturation) where multiple MT nucleation and anchoring factors concentrate forming the poles of the mitotic spindle. Centrosome and centriole size are tightly controlled during the cell cycle and development . C. elegans centrioles are 150 nm long and 100nm wide. Drosophila centrioles vary from 120nm in length in embryos, 180nm in somatic cells to up to 2m in length during spermatogenesis. How these differences and, in particular, how the length of the centriole is regulated in the different cell types is not at all understood. " Cilia formation and function Besides their function in organising the centrosome, centrioles also form the basal bodies at the base of cilia and flagella. Older mother centrioles migrate to the cell cortex and build a transition zone to recruit and anchor the building blocks of the cilia. Interflagelar transport (IFT) modules are assembled and bring the single components to the tip of the growing cilia until a complete and functional cilium is built. Cp110 and Cep97, two centriolar proteins, have been shown to form a cap at the end of the centriole barrel. Absence of Cp110 or Cep97 has been shown to induce cilia formation. How Cp110 and Cep97 regulate centriole length and how it is involved in the conversion from centriole to basal body is not understood. Misregulation of centrosomes affects cell cycle progression and is a hallmark of many tumors Mutations in centrosome and cilia components have been linked to neurological diseases, obesity, left-right asymmetry defects and poly-cystic kidney disease. Thus, the poorly understood mechanisms governing centriole size and its transition from centriole to basal body represent an important open question in cell biology. " Future project I plan to investigate the function of the centriole cap in both Drosophila and C.elegans and focus on Cp110 and its interaction partner Cep97. I identified Cep97 and Cp110 in a genome-wide screen in Drosophila for factors involved in centrosome duplication and maturation. Initial characterisation showed they are bona-fide centriole proteins and localise to the distal end of the centriole barrel in Drosophila. First, I will investigate the function of Cep97 and Cp110 in flies and worms using RNAi depletion and the characterisation of available mutants. Second, GFP-tagged constructs will be made. The dynamic behaviour of the protein and its localisation during development will be tested in different cell types using state-of-the-art microscopy techniques. Finally, pull-down and yeast-2- hybrid as well as RNAi-screening will be developed to identify the constituents of the centriole cap in C.elegans and Drosophila. The proposed experiments will help to explain how eukaryotic cells control the length of centrioles and how the initial stages of cilia formation are controlled. Studying the much more simply built centrioles in the model systems C. elegans and Drosophila will help to understand the molecular function of the recently indentified centriole cap. Furthermore, this analysis will shed light on how mutations in centriole cap proteins in humans result in tumourogenesis and neurological disorders like microcephaly.