Poster abstracts

Poster number 9 submitted by Sana Shaikh

Sarcolipin interaction with SERCA is distinct from phospholamban; only sarcolipin can promote uncoupling of the SERCA pump

Sana A. Shaikh (Department of Physiology and Cell Biology, The Ohio State University, College of Medicine), Sanjaya K. Sahoo (Department of Physiology and Cell Biology, The Ohio State University, College of Medicine), Danesh H. Sopariwala (Department of Physiology and Cell Biology, The Ohio State University, College of Medicine), Naresh C. Bal (Department of Physiology and Cell Biology, The Ohio State University, College of Medicine), Muthu Periasamy (Department of Physiology and Cell Biology, The Ohio State University, College of Medicine)

Abstract:
Sarcolipin (SLN) and Phospholamban (PLB) modulate the activity of the Sarco-Endoplasmic Reticulum Ca²⁺ ATPase (SERCA) pump, in cardiac and skeletal muscle. Recent data suggest that SLN could play a role in muscle thermogenesis by promoting the uncoupling of SERCA ATP hydrolysis activity from its Ca²⁺ transport function. However, the mechanistic details of this uncoupling are unknown. In order to define how the binding of SLN to SERCA promotes uncoupling of SERCA, we compared the interaction of SLN and SERCA1, with that of PLB. We used the homobifunctional cross-linker (1, 6-bismaleimidohexane (BMH)) to detect dynamic protein interaction during the SERCA cycle. We also performed Ca²⁺ uptake and ATP hydrolysis assays to study the effect of SLN on SERCA. Our studies reveal that SLN differs significantly from PLB: 1) SLN primarily affects the V_max of SERCA mediated Ca²⁺ uptake but not the pump affinity for Ca²⁺ 2) SLN can bind to SERCA in the presence of high [Ca²⁺] whereas PLB can only interact with 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. 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 Ca²⁺ causes uncoupling of the SERCA pump and increased heat production.

References:
Lee, A.G. (2002) Curr. Opin. Struct. Biol.12, 547–554
Bal, N. C. et al. (2012) Nat. Med. 18, 1575–1579

Keywords: SERCA, Sarcolipin, Phospholamban