The long-term goal of the proposed studies is to gain a mechanistic understanding of how the cell's microenvironment regulates cytokinesis in complex 3D physiological contexts. Cytokinesis ensures the proper segregation of genetic and cytoplasmic material between daughter cells in the final step of cell division, and is essential fo normal development and tissue homeostasis. Cells interact with their extracellular matrix environment using integrin receptors. There is now compelling evidence that integrins promote the successful completion of cytokinesis; however, the molecular mechanisms are not well understood. Our preliminary data indicate that integrins regulate abscission, the last step in cytokinesis, and point to CHMP4B as a specific target of integrin regulation. Identification of CHMP4B, a component of the ESCRT-III machinery that localizes to the intercellular bridge (or midbody) connecting dividing cells, as a target of integrin regulation overcomes a significant barrier to understanding the mechanistic basis of integrin regulation by identifying a specific molecular target. The goal of Aim 1 is to elucidate the mechanisms by which integrins regulate CHMP4B. Our data also implicate RSK signaling downstream of integrins in the regulation of abscission and demonstrate its localization in midbodies connecting dividing cells.
Aim 2 will identify the mechanisms by which integrin-RSK signaling promotes abscission and determine whether RSK regulates CHMP4B downstream of integrins or by an independent mechanism. Mechanisms will be elucidated in cell culture where integrin activity and signaling can be easily manipulated and specific mechanisms can be easily analyzed, and then the contribution of these mechanisms will be examined in progressively more complex physiological contexts. Mammalian cytokinesis has historically been analyzed using established cell lines in 2D culture; however whether regulatory mechanisms translate to more complex physiological contexts is not known. To address this critical issue, the murine submandibular salivary gland (SMG) will be employed as a model, as our preliminary studies indicate that integrins promote abscission in SMG epithelial cells in 3D culture and during SMG morphogenesis in primary organ culture.
In Aim 3, organ culture and genetic mouse models for conditional, epithelial deletion of a3 and/or a6 integrins will be employed to determine whether integrins and RSK regulate cytokinesis in these complex physiological contexts by mechanisms similar to those identified in cell culture. The proposed studies are significant because they will elucidate the molecular mechanisms by which integrins promote cytokinesis, beginning with specific molecular targets and pathways, and will analyze these mechanisms in increasingly complex physiological contexts. These studies will provide new insight into processes that impact tumorigenesis and development.
Cytokinesis is an important step in cell proliferation, as it ensures the proper separation of genes and other cellular material into two daughter cells. Defects in cytokinesis can lead to cancer and developmental problems. The goal of proposed studies is to understand how a cell's environment can determine cytokinesis success or failure.
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