A High-Turnover Phenotype of Calsequestrin with Elevated Rates of Synthesis and Degradation Replaces Normal Calsequestrin in Heart Failure

Abstract

Left ventricular hypertrophy involves an increase in the size of cardiomyocytes. A central feature of the phenotype is accelerated protein synthesis, but changes are complex, involving increases in protein translation capacity, efficiency, and turnover. We purified calsequestrin (CSQ), a major protein component of the Ca2+ release protein complex, from frozen heart samples of sham control mongrel dogs, or dogs subjected to either tachycardia-induced or microembolism (ischemia)-induced heart failure. We found that the levels of CSQ protein did not change significantly as a result of either of these two treatments. CSQ recovered from control dog heart tissue was, however, fundamentally different from that present in normal dogs. CSQ undergoes both co-translational addition of an N-linked glycan as well as an unusual co-translational phosphorylation of its C-terminus. In control heart tissue, CSQ exhibited its characteristic polymorphic pattern of post-translational demannosylation and dephosphorylation. Failing hearts, however, was marked by large percentages of CSQ molecules (20-40% of total levels) with Man9,8 glycans, reflecting more newly synthesized protein in rough ER. In addition, there was a significant “retrograde shift” in the average glycan processing of CSQ glycans, with longer-lived CSQ molecules replaced with shorter-lived. Such changes indicate a major degradation and biosynthesis (increased turnover) phenotype for CSQ, perhaps representing changes needed to support increased hypertrophic growth. Two phenotypes are also evident using a SDS gel-based analysis. Taken together with our recent findings that CSQ is synthesized around myonuclei and phosphorylated on its C-terminus during its biosynthesis, we propose that qualitative alterations in CSQ metabolism occur in hypertrophic heart. In this proposed model, increased CSQ synthesis leads to increased CSQ in perinuclear cisternae, while increased CSQ degradation prevents buildup of protein in junctional SR cisternae.