Repression of the protein kinase PIM3 by an mTORC1-regulated microRNA

Abstract

The mechanistic target of rapamycin complex 1 (mTORC1) is a central regulator of cell growth that is often aberrantly activated in cancer. However, mTORC1 inhibitors, such as rapamycin, have limited effectiveness as single agent cancer therapies, with feedback mechanisms inherent to the signaling network thought to diminish the anti-tumor effects of mTORC1 inhibition. The goals of this dissertation were to characterize pro-survival effectors activated upon mTORC1 inhibition, and to determine the functional significance of these downstream targets, including relevance to the development of targeted therapies in combination with mTORC1 inhibitors. I identify the repression of protein kinase and proto-oncogene PIM3 downstream of mTORC1 signaling. PIM3 expression is suppressed in cells with loss of the tuberous sclerosis complex (TSC) tumor suppressors, which exhibit growth factor-independent activation of mTORC1, and in the mouse liver upon feeding-induced activation of mTORC1. Inhibition of mTORC1 with rapamycin induces PIM3 transcript and protein levels in a variety of settings. Suppression of PIM3 involves the sterol regulatory element-binding (SREBP) transcription factors SREBP1 and 2, whose processing and mRNA expression are stimulated by mTORC1 signaling. I found that PIM3 repression is mediated by miR-33, an intronic microRNA encoded within the SREBP loci, the expression of which is decreased with rapamycin. I sought to better understand the functional implications of miR-33 induction by mTORC1, and the subsequent induction of PIM3 upon mTORC1 inhibition. Specifically, I show that PIM inhibition in combination with mTOR inhibitors may be a promising therapy in some cancer settings. I also identify several additional mTORC1-regulated miR-33 targets that contribute to cell survival and metabolism, including PIM1, which is closely related to PIM3. Finally, I explore the metabolic changes affected by PIM inhibition, providing an additional rationale for the regulation of PIM3 by mTORC1. Collectively, these studies identify a pro-survival kinase that is activated upon mTORC1 inhibition while highlighting the importance of further characterization of miR-33 targets altered downstream of mTORC1. Our results will guide future studies of mTORC1-regulated microRNAs and pro-survival pathways, with potential implications for the effects of mTORC1 inhibitors in TSC, cancer, and the many other disease settings influenced by aberrant mTORC1 signaling.