The area of microRNA biology is relatively new in our laboratory. However, we are interested in expanding the scope of our research to include microRNAs, since they are major regulators of gene expression but their influence during the stress and proliferative responses is poorly understood. We are particularly well poised to investigate microRNAs because their effects on mRNA stability and translation are similar in many respects to those of RBPs; therefore, we already have the cell systems, methodologies, and expertise in place to study them. To investigate microRNA levels and function in mammalian cells, we employ approaches such as microRNA overexpression (by transfecting microRNA precursors), microRNA reduction (by transfecting antisense microRNAs), and the identification of microRNA-associated mRNAs using biotinylated microRNAs to affinity-purify bound transcripts. We investigate whether microRNAs affect the stability of target mRNAs by measuring the steady-state levels and half-lives of the mRNAs of interest as a function of microRNA abundance. We investigate whether microRNAs affect the translation of target mRNAs by studying the relative assocation of the mRNA with translating polysomes and by quantifying the nascent translation rates of the encoded proteins. We also employ reporter constructs to gain additional insight into these processes. At the beginning of this funding period, we reported (Kim et al., Genes Dev) that HuR reduces c-Myc expression by recruiting let-7 to the c-Myc 3'UTR. Collaborative studies that followed up on the regulation of p16 expression by miR-24 have also identified c-Myc and E2F2 as effectors of miR-24-mediated inhibition of proliferation (Molecular Cell, 2009). Submitted work describes the influence of microRNAs that affect adipogenesis. Ongoing studies are analyzing microRNAs that control expression of other signaling, proliferative, and cancer-related proteins.
| Kim, Hyeon Ho; Kuwano, Yuki; Srikantan, Subramanya et al. (2009) HuR recruits let-7/RISC to repress c-Myc expression. Genes Dev 23:1743-8 |
| Lal, Ashish; Navarro, Francisco; Maher, Christopher A et al. (2009) miR-24 Inhibits cell proliferation by targeting E2F2, MYC, and other cell-cycle genes via binding to ""seedless"" 3'UTR microRNA recognition elements. Mol Cell 35:610-25 |
