Abstract 1468: Posttranscriptional Downregulation Of Mir-1 By The RNA-binding G3BP Is Required For Induction Of eIF4E During Cardiac Hypertrophy
MicroRNAs (miRNAs) are emerging as fundamental posttranscriptional regulators of gene expression. They are differentially regulated during cardiac hypertrophy and failure with major consequences for cellular functions. We have reported earlier that miR-1 was acutely downregulated upon the application of pressure overload to the mouse heart prior to any detectable increase in heart weight. This reduction in miR-1 is essential for the development of hypertrophy. In this study we further investigate the role of miR-1 in hypertrophy and the mechanism involved in its rapid downregulation. Our first hypothesis was that if miR-1 played an essential role in the increase in myocyte mass then it is expected to target genes that are involved in global protein synthesis. Using target prediction software we identified eukaryotic initiation factor 4E (eIF4E) as a target of miR-1. This was confirmed by both protein expression of eIF4E and a luciferase-eIF4E 3′UTR reporter construct in cells reconstituted with miR-1-independent eIF4E gene. The results also show that neutralizing miR-1 downregulation completely inhibited the increase in hypertrophy-induced eIF4E. Overexpression or knockdown of eIF4E in cardiac myocytes is currently underway to quantify its role in hypertrophy and miR-1’s effect. Our second hypothesis was that the rapid downregulation of miR-1 is mediated by a posttranscriptional mechanism, plausibly through an RNA-binding regulatory protein. The Ras-GAP SH3 domain binding protein (G3BP) is an RNA-binding and hydrolyzing protein that is required for myocyte hypertrophy. It binds to and degrades single RNA strands through a specific consensus sequence that we found present in the premature miR-1 (ACCCAUAUGAAC). An adenovirus-mediated overexpression of G3BP in cultured neonatal myocytes elicited a decrease in miR-1 levels (~50%) comparable to the downregulation associated with hypertrophy. On the other hand, silencing G3BP resulted in inhibition of hypertrophy-induced downregulation of miR-1 and the increase in protein synthesis, cell size, and eIF4E. Thus, rapid downregulation of miR-1 occurs via G3BP-induced hydrolysis and is essential for induction of global protein synthesis by releasing eIF4E from its inhibition.