Phenotypic Differences Among Mice with Induced Cardiomyocyte‐Restricted Ablation of Cops5, Cops8, or Both

Abstract

Background- Within a large subset of heart failure, the ubiquitin proteasome system has shown to be inadequate or even causative. The COP9 signalosome (CSN) holocomplex, a vital ubiquitination regulator, directly regulates Cullin-RING ligases (CRLs) via Cullin deneddylation. This holocomplex is formed by 8 unique subunits (COPS1~COPS8), where the deneddylase activity resides in COPS5. Our lab has shown in the past that cardiomyocyte-restricted knockout (CKO) of the Cops8 gene causes cardiomyocyte necroptosis, dilated cardiomyopathy, and shortened lifespan in mice. This study tests whether the protection against necroptosis by Cops8 is deneddylation-dependent or Cops8-specific, possibly alluding to subunit-specific functions independent of the holocomplex. Methods and Results- CKO of Cops8, Cops5, or both in adult mice was achieved using a tamoxifen-inducible Cre-LoxP system. Echocardiography performed 21 days after tamoxifen withdrawal showed no significant difference between MerCreMer (MCM) transgenic and Cops8-floxed/Cops5-floxed control groups. Compared with either control group, all CKO groups developed dilated cardiomyopathy, but the severity in the Cops5-CKO and the Cops8+Cops5 double CKO (dCKO) groups was greater than that in the Cops8-CKO group. Kaplan-Meier survival analyses revealed shortened lifespans for all CKO groups, compared with the MCM group. The post-tamoxifen lifespans of Cops5-CKO and dCKO mice were comparable (median 42 days vs. 42 days; p=0.66) but significantly shorter than that of the Cops8-CKO (81 days, p<0.0001). In Cops5-CKO and dCKO mice, we observed an increased number of cardiomyocytes positive for Evan's blue dye uptake and increased myocardial CD45 proteins compared to Cops8-CKO. All CKO myocardium showed increases in full-length caspase 8 but decreased cleaved caspase 8, indicating suppression of caspase 8 activation by defective CSN. Conclusions (1)Cops5-CKO and dCKO are similarly detrimental but are more so than Cops8-CKO; (2) the primary defect caused by Cops8-CKO results from impaired CSN holocomplex formation and thereby loss of Cullin deneddylation.