Human phospholamban null results in lethal dilated cardiomyopathy revealing a critical difference between mouse and human

Abstract

In human disease and experimental animal models, depressed Ca2+ handling in failing cardiomyocytes is widely attributed to impaired sarcoplasmic reticulum (SR) function. In mice, disruption of the PLN gene encoding phospholamban (PLN) or expression of dominant-negative PLN mutants enhances SR and cardiac function, but effects of PLN mutations in humans are unknown. Here, a T116G point mutation, substituting a termination codon for Leu-39 (L39stop), was identified in two families with hereditary heart failure. The heterozygous individuals exhibited hypertrophy without diminished contractile performance. Strikingly, both individuals homozygous for L39stop developed dilated cardiomyopathy and heart failure, requiring cardiac transplantation at ages 16 and 27. An over 50% reduction in PLN mRNA and no detectable PLN protein were noted in one explanted heart. The expression of recombinant PLN-L39stop in human embryonic kidney (HEK) 293 cells and adult rat cardiomyocytes showed no PLN inhibition of SR Ca2+-ATPase and the virtual absence of stable PLN expression; where PLN was expressed, it was misrouted to the cytosol or plasma membrane. These findings describe a naturally-occurring loss-of-function human PLN mutation (PLN null). In contrast to reported benefits of PLN ablation in mouse heart failure, humans lacking PLN develop lethal dilated cardiomyopathy

Sarcolipin retention in the endoplasmic reticulum depends on its C-terminal RSYQY sequence and its interaction with sarco(endo)plasmic Ca 2 +-ATPases

“…The expression of recombinant Leu39stop cDNA in HEK-293 cells showed the absence of stable PLN protein expression, even with coexpression of stoichiometric amounts of SERCA2a, and no PLN inhibition of SERCA2a activity (24). As well, PLN protein was not detected in immunoblots of a biopsy from the failed heart of a Leu39stop patient (24), consistent with the view that truncated forms of both PLN and SLN are unstable and subject to intracellular degradation. IPs were performed with anti-FLAG antibody, and samples were separated by SDS͞PAGE and subjected to autoradiography.…”

Highly efficient and specific modulation of cardiac calcium homeostasis by adenovector-derived short hairpin RNA targeting phospholamban

“…This appears to be particularly important in humans, as chronic PLB deficiency owing to genomic mutations was associated with cardiomyopathies. [28][29][30] Whereas in mice complete knockout of PLB (as may also be achieved by RNAi) was able to rescue the severe cardiomyopathic phenotype of MLP knockout mice, 31 suggesting that unregulated PLB silencing is appropriate in this species, application in humans most probably requires regulatable RNAi.…”

Sarcolipin and phospholamban as regulators of cardiac sarcoplasmic reticulum Ca2+ ATPase

“…Thus, an alteration in the PLB: SERCA ratio can affect SR Ca 2+ transport. More recently, studies in human suggest that mutations in PLB [43,44] or the absence of PLB [45] can cause far more serious functional consequences, culminating in human heart failure. It is tempting to speculate that, in larger mammals PLB is essential for maintaining heart function, unlike in mouse.…”