Abstract
The E3 ubiquitin ligase, von Hippel–Lindau (VHL), regulates protein expression by polyubiquitination. Although the protein VHL (pVHL) was reported to be involved in the heart function, the underlying mechanism is unclear. Here, we show that pVHL was upregulated in hearts from two types of genetically dilated cardiomyopathy (DCM) mice models. In comparison with the wild-type mouse, both DCM mice models showed a significant reduction in the expression of phospholamban (PLN), a potent inhibitor of sarco(endo)plasmic reticulum Ca2+-ATPase, and enhanced interaction between pVHL and PLN. To clarify whether pVHL is involved in PLN degradation in failing hearts, we used carbonylcyanide m-chlorophenylhydrazone (CCCP), a mitochondrial membrane potential (MMP)-lowering reagent, to mimic the heart failure condition in PLN-expressing HEK293 cells and found that CCCP treatment resulted in PLN degradation and increased interaction between PLN and pVHL. However, these effects were reversed with the addition of N-acetyl-l-cysteine. Furthermore, the co-transfection of VHL and PLN in HEK293 cells decreased PLN expression under oxidative stress, whereas knockdown of VHL increased PLN expression both under normal and oxidative stress conditions. Together, we propose that oxidative stress upregulates pVHL expression to induce PLN degradation in failing hearts.
Highlights
The ubiquitin/proteasome system (UPS) and autophagy/lysosome pathways are the major pathways for intracellular protein degradation and play important roles in protein quality control and various biological processes such as protein folding, signaling transduction, tumorigenesis, molecular trafficking, and clearance [1,2,3]
The expression level of protein VHL (pVHL) is upregulated in response to oxidative stress induced by non-steroidal anti-inflammatory drugs (NSAIDs) [20]
We found that pVHL expression level was upregulated in the heart tissues from the two dilated cardiomyopathy (DCM) mice types (Figure 1a,b)
Summary
The ubiquitin/proteasome system (UPS) and autophagy/lysosome pathways are the major pathways for intracellular protein degradation and play important roles in protein quality control and various biological processes such as protein folding, signaling transduction, tumorigenesis, molecular trafficking, and clearance [1,2,3]. The degradation of HIF-1α by pVHL is known to be regulated through prolyl hydroxylase domain (PHD) proteins [17,18,19]. PVHL is reported to be involved in the heart function by HIF-1α regulation through PHDs [21], the role of pVHL in UPS-mediated protein degradation in the heart is questionable. We demonstrate for the first time that pVHL contributes to the ubiquitination of PLN, thereby inducing its degradation, under oxidative stress conditions in failing hearts (e.g., dilated cardiomyopathy (DCM)). The increase in pVHL-mediated degradation of PLN results in the decreased inhibitory effect of PLN on SERCA2a, possibly leading to impaired SR Ca2+ cycling. Our results suggest that pVHL may act as one of the major E3 ligases that ubiquitinate PLN to induce its degradation under oxidative stress conditions in failing hearts
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