Study of the molecular and functional role of Nek7 and Nek10 in the regulation of cell division associated with cancer

• Grant number: 14/06873-9
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• Start date: November 01, 2014
• End date: October 31, 2017
• Field of knowledge: Biological Sciences - Biochemistry - Molecular Biology
• Agreement: Coordination of Improvement of Higher Education Personnel (CAPES)
• Principal Investigator: Jörg Kobarg
• Grantee: Edmarcia Elisa de Souza
• Host Institution: Instituto de Biologia (IB). Universidade Estadual de Campinas (UNICAMP). Campinas , SP, Brazil
• Associated scholarship(s): 16/07815-8 - Functional and molecular implication of human Nek10 in centrosome function, BE.EP.PD

Abstract

The cell cycle is a complex and well orchestrated process that culminates in cell division, when sets of duplicated chromosomes are partitioned equally between two daughter cells. The mitotic spindle apparatus is composed of central microtubules networks attached to kinetochores and organized from two centrosomes that mediate the proper chromosomes congression. The synergy between the events that drive the spindle and centrosome formation is essential to proper cell division and their deregulation results in hallmark phenotypes of human cancers. The human Neks protein kinases are essential for cell division progression and therefore have been raised attention in cancer therapy. However, the potential of Neks as targets for cancer therapy is relatively unexplored mostly because limited studies regarding their functions have been conducted. Our previous works have been shown that Nek7 and Nek10 are localized at the centrosome, mitotic spindle, and spindle pole during metaphase; central spindle during anaphase; e citoplasmatic bridge and midbody during citokinesis. Furthermore, in yeast two-hybrid studies we showed that Nek7 and Nek10 interact with proteins that contribute to key processes of cell division such as: G2/M transition of mitotic cell cycle, mitotic prometaphase, S phase of mitotic cell cycle, and mitotic cell cycle checkpoint. In this work, we will explore the potential molecular and functional roles of Nek7 and Nek10 in regulating mechanisms of cell division focusing on the centrosome and mitotic spindle functions. To this end, knock down cells and mutant overexpressing wild type, kinase dead and phosphomimetic forms of Nek7 and Nek10 will be generated (step partially completed) and thus assessed for morphological, structural and dynamic phenotypes of cell division using high-resolution fluorescence microscopy and time-lapse image. Furthermore, centrosome purification and microtubule nucleation assays combined with Western Blotting analysis will be performed in order to obtain a direct evidence of Nek7 and Nek10 functions at the centrosome and spindle. This research will greatly contribute to the fundamental understanding of the conceivable regulatory role of Nek7 and Nek10 in regulatting centrosome and spindle during cell division. In summary, besides the relevance of understanding the potential roles of Neks in cell division crucial events for purposes of fundamental research, we expect that the Neks may be indicated as prospective targets for cancer treatment. (AU)