S-nitrosylation of TRIM72 at cysteine 144 is critical for protection against oxidation-induced protein degradation and cell death

Mark J. Kohr, Alicia M. Evangelista, Marcella Ferlito, Charles Steenbergen, Elizabeth Murphy

Research output: Contribution to journalArticlepeer-review

Abstract

Oxidative stress and membrane damage following myocardial ischemia/reperfusion injury are important contributors to cardiomyocyte death and the loss of myocardial function. Our previous study identified cysteine 144 (C144) of tripartite motif-containing protein 72 (TRIM72) as a potential site for S-nitrosylation (SNO). TRIM72 is a cardioprotective membrane repair protein that can be both activated and targeted for degradation by different oxidative modifications. Consistent with the potential regulation of TRIM72 by various oxidative modifications, we found that SNO levels increased at C144 of TRIM72 with ischemic preconditioning. Therefore, to investigate the role of C144 in the regulation of TRIM72 function, we mutated C144 of TRIM72 to a serine residue (TRIM72C144S), and expressed either TRIM72WT or TRIM72C144S in HEK-293 cells, which lack endogenous TRIM72, in order to examine the effect of this mutation on the functional stability of TRIM72 and on cell survival. We hypothesized that SNO of TRIM72 stabilizes the protein, thus allowing for membrane repair and enhanced cell survival. Upon treatment with hydrogen peroxide (H2O2), we found that TRIM72WT levels were decreased, but not TRIM72C144S and this correlated with increased H2O2-induced cell death in TRIM72WT cells. Additionally, we found that treatment with the cardioprotective S-nitrosylating agent S-nitrosoglutathione (GSNO), was able to preserve TRIM72WT protein levels and enhance TRIM72WT-mediated cell survival, but had no effect on TRIM72C144S levels. Consistent with our hypothesis, GSNO was also found to increase SNO levels and inhibit H2O2-induced irreversible oxidation for TRIM72WT without affecting TRIM72C144S. In further support of our hypothesis, GSNO blocked the ischemia/reperfusion-induced decrease in TRIM72 levels and reduced infarct size in a Langendorff-perfused heart model. The results of these studies have important implications for cardioprotection and suggest that SNO of TRIM72 at C144 prevents the oxidation-induced degradation of TRIM72 following oxidative insult, therefore enhancing cardiomyocyte survival.

Original languageEnglish (US)
Pages (from-to)67-74
Number of pages8
JournalJournal of Molecular and Cellular Cardiology
Volume69
DOIs
StatePublished - Apr 2014

Keywords

  • Cardioprotection
  • Cell death
  • Mitsugumin 53
  • Oxidation
  • S-nitrosylation
  • Tripartite motif-containing protein 72

ASJC Scopus subject areas

  • Molecular Biology
  • Cardiology and Cardiovascular Medicine

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