Dr Paolo D'Avino
Many cancer cells present numerical chromosomal abnormalities such as aneuploidy and/or polyploidy. Since the segregation of the genomic material occurs during cell division, an understanding of the mechanisms that control mitosis can unveil some of the processes that promote tumour onset and development. Moreover, one of the hallmarks of cancer is uncontrolled cell proliferation and many cell division regulators are validated targets for the isolation of novel chemotherapeutic drugs for the treatment of cancer pathologies. The goal of our research project is to understand some of the molecular mechanisms that control the last phase of cell division – cytokinesis – and to identify novel gene products that functions in this process in metazoans.
Cleavage furrow formation and ingression during cytokinesis
Cytokinesis ensures the proper distribution of genomic and cytoplasmic material at the end of cell division. This finely orchestrated process requires the co-ordinated action of several proteins that promote a series of sequential events. First, the cell determines the position of the cleavage site through signals generated by the spindle microtubules, which promote the ingression of a cleavage furrow that bisects the dividing cell. Cleavage furrow ingression is driven by the assembly and contraction of actomyosin filaments that often organise into an annular structure known as the contractile ring. The small GTPase RhoA and its downstream effectors control the formation and constriction of this contractile ring. Finally, new membrane, in the form of vesicles transported along microtubules, is inserted at the cleavage site and the two daughter cells become physically separated during abscission. A major focus of the lab is to dissect the signalling pathways downstream of RhoA that control the assembly and constriction of the actomyosin ring. For this reason, we are studying the functions of one of the major RhoA effector during cytokinesis – Citron kinase – in two different model system: the fruitfly Drosophila melanogaster and human tissue culture cells.
Proteomics survey of cell division in Drosophila
The intricate processes of mitosis and cytokinesis are regulated by several protein complexes that mediate spatio-temporally co-ordinated protein phosphorylation and dephosphorylation, proteolytic degradation, and protein folding. Thus a thorough understanding of the mechanics and control of cell division requires the identification of all the individual components of these protein complexes. In collaboration with the groups of Prof. Glover (Department of Genetics, University of Cambridge) and Prof. Laue (Department of Biochemistry, University of Cambridge), we have developed an experimental strategy to isolate protein complexes involved in mitosis and cytokinesis in Drosophila cultured cells. This method involves the tagging of the bait protein with two IgG binding domain of Protein A and the isolation of the tagged bait along with its interacting partners by a single affinity purification step. These isolated complexes can then be analysed by several methods including mass spectrometry and Western blotting. Thanks to this methodology, we have identified partners of more than sixty different proteins involved in different aspects of cell division and my lab is studying some of the most interesting findings from this proteomic survey that might be important for cytokinesis.