Application Of Proteomics In Cell Biology Research

Protein-protein interaction networks orchestrate cellular dynamics and constitute the plasticity underlying cell growth, movement, and specification. Molecular elucidation of protein-protein interaction is essential for understanding the cellular dynamics and plasticity. The completion of genome sequencing project together with the recent development of mass spectrometric technology propel the development of proteomics for resolving complex biological structure. Using proteomics analysis, I have identified a novel ARF6 GTPase activating protein ACAP4 essential for cell migration.During cell division, chromosome segregation is orchestrated by the interaction of spindle microtubules with the centromere. A dramatic reorganization of interpolar microtubules into a highly organized central spindle between the separating chromatids is required for the initiation and execution of cytokinesis. Central spindle organization requires mitotic kinesins and the chromosomal passenger of Aurora B kinase complex with INCENP and survivin. The current working model argues that INCENP targets Aurora B to its points of action while the precise role of survivin is not yet known. My attempt to dissect midbody proteome led to the re-discovery of MKLP2. My functional studies show that human survivin governs Aurora B kinase activity and its relocation to the central spindle using mitotic kinesin MKLP2. We found that survivin interacts with MKLP2 C-terminal tail and disruption of MKLP2-survivin interaction prevents the relocation of Aurora B-INCENP protein complex to the central spindle and caused cytokinesis defects. This MKLP2-mediated, survivin- dependent relocation of the Aurora B protein complex was demonstrated by a set of rescue experiments in which exogenous expression of full-length but not motor-less or tail-less MKLP2 restores the Aurora B kinase complex trafficking and faithful cytokinesis. Surprisingly, survivin-deficiency resulted in minimized Aurora B kinase activity and aberrant targeting of mitotic kinesin MKLP1. The importance of the Aurora B phospho-regulation was validated by correct targeting of ectopic expression of a phosphorylation-mimicking mutant of MKLP1 and faithful completion of cytokinesis. These results indicate that survivin represents a link between the relocation of Aurora B kinase activity to central spindle and completion of cytokinesis. Besides these kinesins and chromosomal passengers, some centrosomal proteins also regulated cytokinesis. Their functional defect would inhibit the completion of cytokinesis. Future experimentation on the spatiotemporal regulation of central spindle dynamics will elucidate the molecular orchestration between chromosome segregation and cytokinesis.