Characterizing the Mechanisms Regulating Myc-induced Transformation

Many current efforts in cancer research focus on understanding the molecular mechanisms driving oncogenesis and to advance molecular diagnostics and targeted therapeutics. The MYC oncoprotein is estimated to be deregulated in over 50% of human cancers, and its deregulation is often associated with aggressive disease and poor patient outcomes. While the ability of MYC to promote cellular transformation is well established, a better understanding of the mechanisms promoting MYC-mediated tumorigenesis is essential. While MYC has been shown to undergo a number of post-translational modifications (PTMs), our current understanding of biological significance of these modifications is largly limited to two phosphorylation sites located in the N-terminal domain of the protein. Our work, therefore, aimed to further our understanding of how PTMs regulate MYC-dependent transformation. To this end we have identified and characterized three novel human cell line models of MYC-dependent transformation: MCF10A, SH-EP Tet21/N-Myc, and LF1/TERT/LT/ST cells. Using a combination of these novel models and classic systems, we have evaluated point mutants of MYC at key serine/threonine and lysine residues for their ability to influence MYC-dependent transformation. Using a six lysine to arginine substitution mutant, we have identified and chacterized six C-terminal lysines to be important for the negative regulation of MYC activity. We have additionally demonstrated for the first time that MYC can undergo SUMOylation at one of the lysines in this region. We further completed a functional and transcriptional characterization of MYC phosphorylation mutants. We have assigned biological significance to previously identified phosphorylation sites through the characterization of two mutants that have increased transformation potential over wild-type MYC. Expression array analysis identified gene expression changes both common to deregulated MYC and unique to the different gain-of-function phosphorylation mutants. Combined, this work has advanced our understanding of several of the mechanisms that may regulate MYC-induced transformation.Ph