Genetic instability is one of the most important hallmarks of cancer. Associations of oncoproteins or tumor suppressors with the process of chromosome segregation provide possible links between carcinogenesis and chromosomal instability. Studies on the function of Rb have been centralized on its role in G1 phase but recently extended to G2/M phases of the cell cycle. Rb interacts with Heclp, through an IxCxE motif, specifically during G2/M phase. Inactivation of hsHeclp by microinjection with anti-Hecl monoclonal antibodies leads to cell death due to abnormal chromosomal segregation. Similarly, inactivation of scHEC1 in budding yeast results in lethality, which can be rescued by hsHEC 1, indicating a highly conserved function for this protein. Heclp appears to be a novel critical protein for M phase progression, and RB can increase the fidelity of chromosome segregation mediated by hsHec lp. Intriguingly, the expression of HEC 1 is highly elevated in most cancer cell lines. Based on these preliminary results, we plan to address the role of Heclp during chromosome segregation in mammalian cells and to explore the possibility of using the Heclp network as a target for developing small molecules that inactivate its function as potential therapeutic candidates. In this application, we proposed four specific aims to address these issues:
Aim 1 : To elucidate the biological significance of the interaction between Hec 1 and Hint 1 at kinetochores and to analyze the role of Heclp and Hintl in spindle checkpoint response.
Aim 2 : To elucidate the biological function of 15A2 and its interaction with Hecl at centrosomes and to analyze the role of Heclp and 15A2 in centrosome functions.
Aim 3 : To examine the regulatory phosphorylation of Hecl by identifying the responsible kinases and to determine roles of the phosphorylated Heclp during chromosome segregation;
and Aim 4 : To explore the Hec 1 networks for potentials as molecular therapeutic targets by identifying small molecules that disrupt the interactions between Heclp and Nek2 as therapeutic candidates. It is expected that elucidation of the function of Heclp will provide useful information in understanding the regulation of chromosome segregation. Moreover, the identified small molecules will not only be a useful tool for dissecting the mitotic machinery, but will have potentials for therapeutic application to treat cancer. ? ?
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