Abstract
Sarco(endo)plasmic reticulum Ca(2+)ATPase (SERCA) pump activity is modulated by phospholamban (PLB) and sarcolipin (SLN) in cardiac and skeletal muscle. Recent data suggest that SLN could play a role in muscle thermogenesis by promoting uncoupling of the SERCA pump (Lee, A.G. (2002) Curr. Opin. Struct. Biol. 12, 547-554 and Bal, N. C., Maurya, S. K., Sopariwala, D. H., Sahoo, S. K., Gupta, S. C., Shaikh, S. A., Pant, M., Rowland, L. A., Bombardier, E., Goonasekera, S. A., Tupling, A. R., Molkentin, J. D., and Periasamy, M. (2012) Nat. Med. 18, 1575-1579), but the mechanistic details are unknown. To better define how binding of SLN to SERCA promotes uncoupling of SERCA, we compared SLN and SERCA1 interaction with that of PLB in detail. The homo-bifunctional cross-linker (1,6-bismaleimidohexane) was employed to detect dynamic protein interaction during the SERCA cycle. Our studies reveal that SLN differs significantly from PLB: 1) SLN primarily affects the Vmax of SERCA-mediated Ca(2+) uptake but not the pump affinity for Ca(2+); 2) SLN can bind to SERCA in the presence of high Ca(2+), but PLB can only interact to the ATP-bound Ca(2+)-free E2 state; and 3) unlike PLB, SLN interacts with SERCA throughout the kinetic cycle and promotes uncoupling of the SERCA pump. Using SERCA transmembrane mutants, we additionally show that PLB and SLN can bind to the same groove but interact with a different set of residues on SERCA. These data collectively suggest that SLN is functionally distinct from PLB; its ability to interact with SERCA in the presence of Ca(2+) causes uncoupling of the SERCA pump and increased heat production.
Highlights
Sarcolipin and phospholamban, the regulators of sarco(endo)plasmic reticulum Ca2ϩATPase (SERCA), are differentially expressed in muscle
Our studies reveal that SLN differs significantly from PLB: 1) SLN primarily affects the Vmax of SERCA-mediated Ca2؉ uptake but not the pump affinity for Ca2؉; 2) SLN can bind to SERCA in the presence of high Ca2؉, but PLB can only interact to the ATP-bound Ca2؉-free E2 state; and 3) unlike PLB, SLN interacts with SERCA throughout the kinetic cycle and promotes uncoupling of the SERCA pump
We investigated whether PLB plays a role in muscle thermogenesis using PLBϪ/Ϫ mice
Summary
Sarcolipin and phospholamban, the regulators of SERCA, are differentially expressed in muscle. Mall et al [28] showed that SLN binding to SERCA promotes uncoupling of the SERCA pump and slippage of Ca2ϩ into the cytoplasm instead of the SR lumen These studies suggested that SLN binding with SERCA can increase ATP hydrolysis and heat production and could contribute to muscle thermogenesis [29, 30]. SLNϪ/Ϫ mice became significantly obese when fed on high fat diet, whereas WT mice were less obese and significantly up-regulated SLN expression [31] These data suggested that the muscle-based SLN-SERCA interaction contributes to heat production and energy expenditure. These findings provide new insight into the molecular basis of SLNSERCA interaction and highlight that SLN alone is responsible for muscle thermogenesis
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