No abstract

Activating mutations of c-kit at codon 816 (Asp 816 ) have been identi®ed in variety of malignancies, including acute myeloid leukemia (AML), mastocytosis and germ cell tumors. The mutant c-Kit receptor confers cytokine independence and induces tumorigenicity. However, the molecular mechanisms, particularly the changes in the signal transduction pathways, responsible for these biological e ects induced by mutant c-Kit are largely unde®ned. Using the human embryonic kidney cell line, 293, we show in the current report that constitutive activation of STAT3 and STAT1 is associated with

IntroductionWe have shown that NF‐κB regulates the expression of over 200 genes in late ischemic preconditioning (IPC) and its activation is necessary for the cardioprotection of late IPC. We have also demonstrated that Hsp70.3 is one of these NF‐κB regulated genes whose expression is necessary to elicit late IPC protection.ResultsUsing a novel non‐viral DNA delivery agent to apply transcription factor decoys to the in vivo murine heart, we show that, in addition to NF‐κB, transcriptional blockade of AP‐1, or STAT3 prior to IPC abrogates the late phase of cardioprotection. We hypothesize that NF‐κB, AP‐1, and STAT3 co‐regulate a common subset of genes, including Hsp70.3, that are critical for late phase IPC cardioprotection. Results using in vitro reporter assays demonstrate coordinated regulation of the Hsp70.3 promoter by NF‐κB, AP‐1, and STAT‐3. We also used PCR based gene microarrays to examine in vivo expression changes of 238 previously identified NF‐κB‐dependent genes under conditions of AP‐1 and STAT3 inhibition after IPC to identity additional genes co‐regulated by this trio of IPC requisite transcription factors.ConclusionsThe set of genes whose expression is co‐dependent on multiple cardioprotective transcription factors after IPC are likely to underlie common critical pathways of cardioprotection and may represent novel therapeutic targets for the treatment of ischemic heart disease.This work was supported by NIH grant R01 HL63034 (WKJ).

No abstract

IntroductionWe showed that the Hsp70.3 isoform is induced by NF‐κB after late ischemic preconditioning (IPC) and contributes to cardioprotection. However, the nearly identical Hsp70.1 isoform does not contribute to IPC induced cardioprotection. We propose that the functional differences between the twoHSP70s are due to post‐transcriptional regulation involving the 3' UTRs, which are divergent.ResultsThough NF‐κB inhibition completely reduces protein expression, it only partially reduces Hsp70.3 mRNA levels, showing that Hsp70.3 is regulated by both pre‐ and post‐transcriptional mechanisms after IPC. We hypothesized that an IPC stimulus reduces expression of Hsp70.3‐suppressing miRNA(s) allowing for post‐transcriptional stabilization of Hsp70.3 mRNA and increased Hsp70.3 protein levels. We assessed in vivo levels of 356 miRNAs in a mouse model of IPC, to identify miRNAs that are; 1) downregulated after IPC, and 2) predicted to target Hsp70.3. We identified miR‐378* as the only miRNA of those surveyed to meet these criteria and demonstrated functional inhibition of Hsp70.3 by miR‐378*.DiscussionThis work demonstrates post‐transcriptional (via miRNA) regulation of Hsp70.3. It appears that coordinated transcriptional upregulation (promoter) as well as suppression of targeting miRNAs are involved in increasing Hsp70.3 protein expression after IPC. This work was supported by the American Society of Pharmacology and Experimental Therapeutics (ASPET) with funds generously provided by ASPET and the University of Cincinnati.