Inadequate ubiquitination-proteasome coupling contributes to myocardial ischemia-reperfusion injury
Chengjun Hu, Yihao Tian, Hongxin Xu, Bo Pan, Erin M. Terpstra, Penglong Wu, Hongmin Wang, Faqian Li, Jinbao Liu, Xuejun Wang
Chengjun Hu, Yihao Tian, Hongxin Xu, Bo Pan, Erin M. Terpstra, Penglong Wu, Hongmin Wang, Faqian Li, Jinbao Liu, Xuejun Wang
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Research Article Cardiology

Inadequate ubiquitination-proteasome coupling contributes to myocardial ischemia-reperfusion injury

Abstract

The ubiquitin-proteasome system (UPS) degrades a protein molecule via 2 main steps: ubiquitination and proteasomal degradation. Extraproteasomal ubiquitin receptors are thought to couple the 2 steps, but this proposition has not been tested in vivo with vertebrates. More importantly, impaired UPS performance plays a major role in cardiac pathogenesis, including myocardial ischemia-reperfusion injury (IRI), but the molecular basis of UPS impairment remains poorly understood. Ubiquilin1 is a bona fide extraproteasomal ubiquitin receptor. Here, we report that mice with a cardiomyocyte-restricted knockout of Ubiquilin1 (Ubqln1-CKO mice) accumulated a surrogate UPS substrate (GFPdgn) and increased myocardial ubiquitinated proteins without altering proteasome activities, resulting in late-onset cardiomyopathy and a markedly shortened life span. When subject to regional myocardial ischemia-reperfusion, young Ubqln1-CKO mice showed substantially exacerbated cardiac malfunction and enlarged infarct size, and conversely, mice with transgenic Ubqln1 overexpression displayed attenuated IRI. Furthermore, Ubqln1 overexpression facilitated proteasomal degradation of oxidized proteins and the degradation of a UPS surrogate substrate in cultured cardiomyocytes without increasing autophagic flux. These findings demonstrate that Ubiquilin1 is essential to cardiac ubiquitination-proteasome coupling and that an inadequacy in the coupling represents a major pathogenic factor for myocardial IRI; therefore, strategies to strengthen coupling have the potential to reduce IRI.

Authors

Chengjun Hu, Yihao Tian, Hongxin Xu, Bo Pan, Erin M. Terpstra, Penglong Wu, Hongmin Wang, Faqian Li, Jinbao Liu, Xuejun Wang

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Figure 9

Ubqln1 overexpression promotes proteasomal degradation of oxidized proteins in cultured NRVMs.

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Ubqln1 overexpression promotes proteasomal degradation of oxidized prote...
Twenty-four hours after plating, NRVMs were infected with Ad-Ubqln1 or Ad–β-Gal. Further experiments were initiated at 48 hours after the adenoviral infection. (A and B) Representative images of Western blot analyses for DNP-derivatized protein carbonyls. The Ubqln1 overexpressing and control NRVMs were treated with H2O2 at the indicated doses. The cells were harvested at either 15 minutes (A) or 3 hours (B) after the H2O2 treatment for DNPH derivatization to measure the level of protein carbonyls. D/G, DNP/GAPDH. Representatives of 3 biological repeats are shown. (C and D) Proteasome inhibition attenuates the Ubqln1 overexpression-induced reduction of oxidized proteins. The cultured NRVMs were treated with proteasome inhibitor bortezomib (BZM, +) or vehicle control (saline, –) at 10 minutes before H2O2 treatment was initiated. Three hours later, cells were harvested for DNPH derivatization. Representative images (C) and pooled densitometry data from 3 biological repeats (D) are shown. *P < 0.05 vs. the H2O2 group; 1-way ANOVA followed by Tukey’s test.