A cell's transition between active proliferation and quiescence (reversible cell cycle arrest) is a cellular event of fundamental importance. Failures of this process cause a wide range of pathologies, including cancer. The long-term goal of this research program is to elucidate the mechanisms and pathways regulating these transitions and to test what role these mechanisms play in tumorigenesis. Recent studies have demonstrated the importance of microRNAs (miRNA) in a wide variety of cellular processes including cancer. The let-7 family of miRNAs has been specifically associated with tumorigenesis, as low let-7 levels have been found in multiple tumors. We have found that let-7b is up-regulated upon induction of quiescence. let-7b overexpression in normal primary fibroblasts results in a decrease in cell number and an increase in the fraction of cells in the G2/M cell cycle phase, let-7 overexpression also results in down-regulation of proteins involved in cell cycle and proteasome- mediated protein degradation, such as Cdc34 (an E2 ubiquitin conjugating enzyme). Cdc34 down-regulation results, in turn, in partial stabilization of Wee1 kinase (an inhibitor of G2/M progression). let-7b up-regulation may help maintain quiescence, in part, by down-regulating proteasome-mediated protein degradation pathway. Low let-7 levels may result in high levels of Cdc34, leading to more rapid proteolysis of cell cycle proteins and ultimately making cells prone to chromosomal instability, with resultant cancerous growth. To explore this hypothesis, we propose: (1) to determine the functional effects of let-7 on proteasome-mediated protein degradation and quiescent cell viability by manipulating the levels of let-7b in primary fibroblasts and monitoring protein degradation and cell viability in proliferating and quiescent cells;(2) to determine whether low let-7b levels contribute to genomic instability and to decipher the possible molecular mechanism by which let-7 affects radiosensitivity;and (3) to determine the possible mechanism by which let-7b is differentially regulated in proliferating and quiescent fibroblasts. Deciphering the mechanisms by which specific let-7 downstream targets control the cell cycle and quiescence may lead to better strategies for controlling cell division and may provide insight into the unrestricted, pathological division characteristic of cancer cells

National Institute of Health (NIH)
National Cancer Institute (NCI)
Research Scientist Development Award - Research & Training (K01)
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Subcommittee G - Education (NCI)
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Ojeifo, John O
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Princeton University
Schools of Arts and Sciences
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Legesse-Miller, Aster; Raitman, Irene; Haley, Erin M et al. (2012) Quiescent fibroblasts are protected from proteasome inhibition-mediated toxicity. Mol Biol Cell 23:3566-81
Wang, David J; Legesse-Miller, Aster; Johnson, Elizabeth L et al. (2012) Regulation of the let-7a-3 promoter by NF-?B. PLoS One 7:e31240
Suh, Eric J; Remillard, Matthew Y; Legesse-Miller, Aster et al. (2012) A microRNA network regulates proliferative timing and extracellular matrix synthesis during cellular quiescence in fibroblasts. Genome Biol 13:R121