The ubiquitin-proteasome system (UPS) is essential for protein degradation and plays critical roles in myocardial ischemia/reperfusion (MI/R) injuries. Previous studies have demonstrated that the soluble receptor for advanced glycation end-product (sRAGE) inhibited MI/R-induced apoptosis by upregulating proteasome subunits. However, the mechanism remains unknown. An MI/R model was established by left anterior descending (LAD) coronary artery ligation in mice. Recombinant sRAGE protein or saline was injected intramyocardially with or without neutralizing interferon-gamma (IFN-gamma) antibody injected intraperitoneally before ligation. In cardiomyocytes, ischemia was simulated with "ischemia buffer" and sRAGE was overexpressed by adenovirus. Adenovirus expressing the interference RNA of beta 5i was used to knockdown beta 5i in cardiomyocytes. IFN-gamma was induced by sRAGE both in sham and MI/R mice. Blockade of IFN-gamma using IFN-gamma antibody abolished the rescue effects of sRAGE for cardiac dysfunction, infarct size and apoptosis provoked by MI/R. Blockade of IFN-gamma reversed the upregulation of beta 1i and beta 5i expression induced by sRAGE during MI/R in heart, accompanied by decreasing chymotrypsin-like proteasome activity. In addition, IFN-gamma antibody abolished the suppressing effect of sRAGE on MI/R-induced p38 and c-Jun N-terminal kinase (JNK) activation, as well as p53 expression, both in vivo and in vitro. However, knockdown of beta 5i abolished the antiapoptosis effect of sRAGE during hypoxia/reoxygenation (H/R) in vitro, accompanied by decreased degradation of p53. Our data suggest a novel mechanism for sRAGE in preventing MI/R-induced apoptosis in heart: sRAGE inhibits MI/R-induced apoptosis in cardiomyocytes by degrading p53 by beta 5i subunit that is increased via upregulation of IFN-gamma.