Riabowol, Karl T.Bose, Pinaki2017-12-182017-12-182010Bose, P. (2010). Ing1 protein function in apoptosis and senescence (Doctoral thesis, University of Calgary, Calgary, Canada). Retrieved from https://prism.ucalgary.ca. doi:10.11575/PRISM/4693http://hdl.handle.net/1880/105694Bibliography: p. 214-232Some pages are in colour.ING proteins act as readers and writers of the histone epigenetic code, affecting DNA repair, cellular senescence and apoptosis. My doctoral research focused upon understanding the molecular mechanisms underlying the involvement of ING1 in stress signaling. My work began with elucidating novel interacting partners of ING proteins using a cross-species (yeast, fly, and human) bioinformatics-based approach. We identified 381 proteins in a yeast interactome analysis that interacted with ING and also had human counterparts. I then biochemically confirmed the validity of the screen, showing that ING1 interacts with three proteins involved in stress signaling: p38MAPK, MEKK4 and RAD50. Our bioinformatics screen indicated that ING proteins in yeast interact with several mitochondrial proteins. Given that ING1 and p53 can functionally interact and that p53 has a transcription-independent role in apoptosis in the mitochondria, I asked if ING1 might also function outside the nucleus. We found that ING1 translocates to the mitochondria in primary fibroblasts and in established epithelial cell lines in response to apoptosis-inducing stimuli, independent of cellular p53 status. I also determined that endogenous ING1 specifically interacts with the pro-apoptotic BCL2 machinery in the mitochondria, suggesting a model in which the ING1-BAX interaction promotes mitochondrial membrane permeability, thereby promoting apoptosis. Since cellular aging is widely thought to represent a form of telomere­induced stress, I also investigated if ING1 interacted with the telomeric protein TRF2 that is involved in DNA damage and repair pathways. We found that, indeed, endogenous ING1 protein interacted with TRF2. Interestingly, TRF2 protein levels were also seen to decrease in senescent primary fibroblasts without a concomitant decrease in TRF2 mRNA levels. Further, our data provides evidence for the fact that telomere binding of TRF2 is required for its stability thereby indicating that a decrease in TRF2 levels might affect the induction of senescence caused by telomere attrition. Lastly, I characterized the DNA damage response in young versus senescent fibroblasts in which ING1 levels are altered, using 53BP1 focus formation as a surrogate marker for DNA damage. I found that decreased ability of senescent cells to process DNA damage foci correlates well with the loss of ING1 b during senescence, consistent with a role for ING1 bin DNA repair.xvi, 232 leaves : ill. ; 30 cm.engUniversity of Calgary graduate students retain copyright ownership and moral rights for their thesis. You may use this material in any way that is permitted by the Copyright Act or through licensing that has been assigned to the document. For uses that are not allowable under copyright legislation or licensing, you are required to seek permission.doctoral thesis10.11575/PRISM/4693