Abstract 21007: Post-Transcriptional Regulation of Hsp70.3 Expression in Late Ischemic Preconditioning by Alternative Polyadenylation
Introduction: Recent results from our lab have demonstrated that NF-κB-dependent expression of heat shock protein 70.3 (Hsp70.3) is required for late ischemic preconditioning (IPC). We have also shown that Hsp70.3 expression is subject to post-transcriptional regulation mediated by microRNA (miRNA) interactions at the 3′-UTR. Upon investigation of the 3′-UTR of the Hsp70.3 mRNA sequence, it was found that multiple polyadenylation signals are predicted to exist within this region. Therefore, we hypothesized that alternative polyadenylation of the Hsp70.3 mRNA transcript, in coordination with miRNA regulation, plays a key regulatory role in Hsp70.3 expression in the myocardium following late IPC.
Results: We utilized rapid amplification of cDNA ends (RACE) to assess the endogenous population of Hsp70.3 transcript sizes and found that four distinct Hsp70.3 transcripts exist as a result of alternative polyadenylation (polyA). We found two predominant populations of alternatively polyadenylated Hsp70.3 transcript that represented the result of polyadenylation at one of two sites on either side of the site for a miRNA that we have shown regulates Hsp70.3 expression. Quantitative analysis of polyA site utilization using QRT-PCR revealed a significant increase in Hsp70.3 mRNA transcripts with shortened 3′-UTRs following an IPC stimulus. Furthermore, this IPC induced shortening of the Hsp70.3 transcript through alternative polyadenylation leads to the removal of a regulatory miRNA binding site.
Discussion: Our results indicate for the first time that alternative polyadenylation plays a role in post-transcriptional regulation of cardioprotective gene programs following a late IPC stimulus. In addition, it appears that these processes may be coordinated with miRNA-mediated post-transcriptional regulation. Investigations are currently ongoing to determine the mechanisms mediating polyadenylation site selection of Hsp70.3 as well as other key mRNA transcripts following late IPC.
- © 2010 by American Heart Association, Inc.