Sarcolipin Research Articles

Mapping of Sarcolipin Conformational States along the Enzymatic Pathway of SERCA

Sarcolipin (SLN), a 31 amino acid transmembrane peptide, binds to the Sarco(endo)plasmic Reticulum Ca2+ ATPase (SERCA) decreasing its apparent Ca2+ affinity. SERCA pumps Ca2+ from the cytoplasm of muscle cells into the sarcoplasmic reticulum (SR) using energy derived from ATP hydrolysis. This re-establishes the Ca2+ gradients needed for normal muscle function. Phosphorylation of SLN relieves the inhibition of SERCA and may be involved in the beta-adrenergic response. Although a significant amount is known about SERCA, and many of the conformations throughout its transport cycle have been crystallized, the mechanism of how SLN decreases SERCA's Ca2+ affinity and alters the coupling between ATP hydrolysis and Ca2+ transport is not well understood. Using solid-state nuclear magnetic resonance spectroscopy and functional assays, we investigated the differences in the interaction of phosphorylated SLN with SERCA compared to non-phosphorylated SLN with SERCA. Previous work in our lab demonstrates that the topology of SLN in complex with SERCA changes depending on SERCA's conformation. As such, continuing work involves mapping the structural changes of phosphorylated SLN with the various conformations of SERCA. Overall, gaining a better understanding of how SERCA is regulated will add in the development of therapies for diseases resulting from improper calcium cycling.

Allosteric Regulation of the Sarcoplasmic Reticulum Ca2+-Atpase by Phospholamban and Sarcolipin using Solid-State NMR Spectroscopy

The membrane protein complexes between the sarcoplasmic reticulum Ca2+-ATPase (SERCA) and phospholamban (PLN) or sarcolipin (SLN) control Ca2+ transport in cardiomyocytes, thereby modulating cardiac muscle contractility. Both PLN and SLN are phosphorylated upon β-adrenergic-stimulated phosphorylation and up-regulate the ATPase via an unknown mechanism. Using solid-state NMR spectroscopy, we mapped the interactions between SERCA and both PLN and SLN in membrane bilayers. We found that the allosteric regulation of the ATPase depends on the conformational equilibria of these two endogenous regulators that maintain SERCA's apparent Ca2+ affinity within a physiological window. Here, we present new regulatory models for both SLN and PLN that represent a paradigm-shift in our understanding of SERCA function. Our data suggests new strategies for designing innovative therapeutic approaches to enhance cardiac muscle contractility.

Cold adaptation overrides developmental regulation of sarcolipin expression in mice skeletal muscle: SOS for muscle-based thermogenesis?

Neonatal mice have a greater thermogenic need than adult mice and may require additional means of heat production, other than the established mechanism of brown adipose tissue (BAT). We and others recently discovered a novel mediator of skeletal muscle-based thermogenesis called sarcolipin (SLN) that acts by uncoupling sarcoendoplasmic reticulum Ca(2+)-ATPase (SERCA). In addition, we have shown that SLN expression is downregulated during neonatal development in rats. In this study we probed two questions: (1) is SLN expression developmentally regulated in neonatal mice?; and (2) if so, will cold adaptation override this? Our data show that SLN expression is higher during early neonatal stages and is gradually downregulated in fast twitch skeletal muscles. Interestingly, we demonstrate that cold acclimation of neonatal mice can prevent downregulation of SLN expression. This observation suggests that SLN-mediated thermogenesis can be recruited to a greater extent during extreme physiological need, in addition to BAT.

S-Palmitoylation and S-Oleoylation of Rabbit and Pig Sarcolipin

Sarcolipin (SLN) is a regulatory peptide present in sarcoplasmic reticulum (SR) from skeletal muscle of animals. We find that native rabbit SLN is modified by a fatty acid anchor on Cys-9 with a palmitic acid in about 60% and, surprisingly, an oleic acid in the remaining 40%. SLN used for co-crystallization with SERCA1a (Winther, A. M., Bublitz, M., Karlsen, J. L., Moller, J. V., Hansen, J. B., Nissen, P., and Buch-Pedersen, M. J. (2013) Nature 495, 265-2691; Ref. 1) is also palmitoylated/oleoylated, but is not visible in crystal structures, probably due to disorder. Treatment with 1 m hydroxylamine for 1 h removes the fatty acids from a majority of the SLN pool. This treatment did not modify the SERCA1a affinity for Ca(2+) but increased the Ca(2+)-dependent ATPase activity of SR membranes indicating that the S-acylation of SLN or of other proteins is required for this effect on SERCA1a. Pig SLN is also fully palmitoylated/oleoylated on its Cys-9 residue, but in a reverse ratio of about 40/60. An alignment of 67 SLN sequences from the protein databases shows that 19 of them contain a cysteine and the rest a phenylalanine at position 9. Based on a cladogram, we postulate that the mutation from phenylalanine to cysteine in some species is the result of an evolutionary convergence. We suggest that, besides phosphorylation, S-acylation/deacylation also regulates SLN activity.

The role of skeletal-muscle-based thermogenic mechanisms in vertebrate endothermy.

Thermogenesis is one of the most important homeostatic mechanisms that evolved during vertebrate evolution. Despite its importance for the survival of the organism, the mechanistic details behind various thermogenic processes remain incompletely understood. Although heat production from muscle has long been recognized as a thermogenic mechanism, whether muscle can produce heat independently of contraction remains controversial. Studies in birds and mammals suggest that skeletal muscle can be an important site of non-shivering thermogenesis (NST) and can be recruited during cold adaptation, although unequivocal evidence is lacking. Much research on thermogenesis during the last two decades has been focused on brown adipose tissue (BAT). These studies clearly implicate BAT as an important site of NST in mammals, in particular in newborns and rodents. However, BAT is either absent, as in birds and pigs, or is only a minor component, as in adult large mammals including humans, bringing into question the BAT-centric view of thermogenesis. This review focuses on the evolution and emergence of various thermogenic mechanisms in vertebrates from fish to man. A careful analysis of the existing data reveals that muscle was the earliest facultative thermogenic organ to emerge in vertebrates, long before the appearance of BAT in eutherian mammals. Additionally, these studies suggest that muscle-based thermogenesis is the dominant mechanism of heat production in many species including birds, marsupials, and certain mammals where BAT-mediated thermogenesis is absent or limited. We discuss the relevance of our recent findings showing that uncoupling of sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA) by sarcolipin (SLN), resulting in futile cycling and increased heat production, could be the basis for NST in skeletal muscle. The overall goal of this review is to highlight the role of skeletal muscle as a thermogenic organ and provide a balanced view of thermogenesis in vertebrates.

Measuring Ca2+-dependent Ca2+-uptake activity in the mouse heart.

The apparent Ca(2+) affinity of the isoforms of the sarco/endoplasmic reticulum Ca(2+) ATPase SERCA2 is controlled primarily by two proteins, phospholamban (PLB) and sarcolipin (SLN). The rate of ATP-driven Ca(2+) uptake into sarcoplasmic reticulum (SR)-derived vesicles can be monitored by a technique in which the net uptake of (45)Ca(2+) in the form of an intravesicular calcium oxalate precipitate is recorded. Here, we present details of a modification of such a protocol for determining the apparent Ca(2+) affinity of the Ca(2+) pump, and its control by various regulators, in crude homogenates of mouse heart.

Identification and purification of human induced pluripotent stem cell-derived atrial-like cardiomyocytes based on sarcolipin expression.

The use of human stem cell-derived cardiomyocytes to study atrial biology and disease has been restricted by the lack of a reliable method for stem cell-derived atrial cell labeling and purification. The goal of this study was to generate an atrial-specific reporter construct to identify and purify human stem cell-derived atrial-like cardiomyocytes. We have created a bacterial artificial chromosome (BAC) reporter construct in which fluorescence is driven by expression of the atrial-specific gene sarcolipin (SLN). When purified using flow cytometry, cells with high fluorescence specifically express atrial genes and display functional calcium handling and electrophysiological properties consistent with atrial cardiomyocytes. Our data indicate that SLN can be used as a marker to successfully monitor and isolate hiPSC-derived atrial-like cardiomyocytes. These purified cells may find many applications, including in the study of atrial-specific pathologies and chamber-specific lineage development.

Sarcolipin provides a novel muscle-based mechanism for adaptive thermogenesis.

The sarco(endo)plasmic reticulum Ca-ATPase (SERCA) transports Ca into the sarcoplasmic reticulum lumen and contributes significantly to skeletal muscle metabolic rate. Sarcolipin (SLN) has been shown recently to uncouple Ca transport from adenosine triphosphate hydrolysis by SERCA. We have hypothesized that SLN provides a novel mechanism of adaptive thermogenesis within skeletal muscle and protects against diet-induced obesity.

Sarcolipin overexpression in mice increases their endurance exercise capacity (1162.2)

Sarcolipin (SLN) is a transmembrane protein that interacts with Sarco(endo)plasmic Reticulum Calcium ATPase (SERCA) and is expressed in cardiac and skeletal muscle. SLN is known to inhibit Vmax of SERCA, but its role in muscle physiology is yet to be fully understood. We have shed some light on this front, by showing that SLN knock out (SLN KO) mice are highly sensitive to acute cold and are prone to develop obesity when fed a high fat diet. Moreover, we showed that the SLN KO mice can be rescued from hypothermia by over expression of the SLN transgene; suggesting that SLN is essential for heat generation and increasing metabolic rate. The aim of this study was to further explore how SLN affects muscle function and metabolism; by the use of SLN over expression (SLN OE) mouse model, which expresses SLN in all skeletal muscles up to 10 fold higher than in wild type (WT) mice. Here, we show that SLN OE mice have a higher basal metabolic rate (by indirect calorimetry) than WT mice. The SLN OE mice also have a significantly higher endurance capacity (treadmill running) than their WT littermates. Isolated muscle studies using Tissue Oxygen bath system, show that SLN OE muscle has similar force frequency response as WT muscle, but is more resistant to develop fatigue and consumes more oxygen. Analysis of muscle oxidative metabolic markers and oxygraph measurements using isolated mitochondria; suggest an increased oxidative capacity in the SLN OE muscle compared to WT. Although skeletal muscle can increase metabolic rate up to hundred fold, a decline in the rate of ATP production is a major reason for muscle fatigue during prolonged physical activity. Hence, these results indicate that SLN enhances muscle’s ability to hydrolyze substrates and generate ATP in times of increased energy demand, which in turn increases endurance capacity.Grant Funding Source: Supported by NIH

Sarcolipin orchestrates calcium signaling to regulate muscle metabolism (1162.1)

Sarcolipin(SLN) is 31 amino acid long protein that regulates the Sarcoplasmic reticulum calcium ATPase (SERCA) activity in skeletal and cardiac muscle. Recently, our lab identified that SLN plays an important role in muscle‐based thermogenesis. Mice lacking SLN are cold sensitive and susceptible to diet induced obesity. Interestingly SLN is induced in skeletal muscle during neonatal development, high fat diet, muscle dystrophies, and inflammation. Since these are the states of increased oxidative metabolism in skeletal muscle we hypothesize that SLN is recruited to maintain this oxidative phenotype. Here we show that SLN overexpressing muscle fibers have high resting calcium and increased recruitment of calcium mediated signaling when there is increased demand. To determine the mechanism regulating SLN induction and downstream signaling we made use of C2C12 cells. We show that inhibiting the calcium signaling pathways regulated by calcineurin downregulates the expression of SLN. Based on our data we propose that SLN forms a feedback loop with calcium mediated signaling pathways and plays an important role to support the oxidative metabolism in skeletal muscle.Grant Funding Source: Supported by NIH, MDA

Sarcolipin interaction with SERCA is distinct from Phospholamban; only Sarcolipin can promote uncoupling of the SERCA pump (LB244)

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 SLN promotes uncoupling of SERCA, we compared the interaction of SLN and SERCA1, with that of PLB. We used the 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 1) SLN affects the Vmax of SERCA 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 different residues on SERCA. These data 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.Grant Funding Source: Supported by NIH and AHA

Loss of SLN‐mediated thermogenesis is compensated by increased brown adipose tissue activity (1105.4)

Thermogenic processes are critical contributors to body temperature maintenance and, when activated, are major components of metabolism. Elucidating the mechanisms that contribute to thermogenesis is of increasing importance as they can provide targets to combat the rising obesity epidemic. We recently identified Sarcolipin (SLN), a regulator of the sarcoplasmic reticulum Ca2+ ATPase (SERCA) in skeletal muscle, as a significant contributor to metabolism and cold‐induced thermogenesis during an acute cold challenge. However, acute cold exposure does not allow for a detailed study of the physiological and molecular mechanisms involved. Thus, to further detail the role of SLN in thermogenesis, we challenged wild‐type (WT) and SLN‐knockout (SLN‐KO) mice to a long‐term cold exposure. SLN‐KO mice can adapt to gradual cold exposure and maintain normal core body temperature. However, SLN‐KO mice exhibit increased oxygen consumption compared to WT littermates and have greater physical activity and food intake. Histological examination of brown adipose (BAT) and inguinal fat depots showed increased depletion of fat stores from cold‐adapted SLN‐KO mice, and protein analysis showed increased UCP1 expression. These data show that SLN‐KO mice must significantly increase BAT activity and browning of white fat in order to compensate for the loss of SLN‐mediated thermogenesis. Increased physical activity, food intake, and elevated metabolic rate suggest a reliance on less efficient thermogenic mechanisms. Thus, an absence of SLN increases the energetic cost of survival in a cold environment. These data demonstrate that a combination of BAT and SLN‐based thermogenesis is necessary for survival in a cold environment and are integral components of energetic homeostasis.

Sarcolipin Regulation of SERCA is Distinct from Phospholamban

Sarco(endo)plasmic reticulum Ca(2+)ATPase (SERCA) pump activity is modulated by phospholamban (PLB) and sarcolipin (SLN) in cardiac and skeletal muscle. Our recent data suggest that SLN plays a role in muscle thermogenesis by promoting uncoupling of the SERCA pump (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 not PLB to SERCA promotes uncoupling of SERCA, we compared SLN and SERCA1 interaction with that of PLB in detail. We used chemical cross-linking to study the dynamic protein-protein interaction. 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 Ca2+ causes uncoupling of the SERCA pump and increased heat production.

Solid-State NMR Structures of Phospholamban or Sarcolipin Bound to Calcium ATPase (SERCA) Reveal the Mode of Inhibition

Sarco(endo)plasmic reticulum calcium ATPase (SERCA) transports Ca2+ ions against a concentration gradient from cytoplasm to sarcoplasmic reticulum, initiating muscle relaxation. SERCA activity is regulated by two single-span membrane proteins: phospholamban (PLN) or sarcolipin (SLN). Although the recent crystal structures offered a prime view of these complexes (SERCA/SLN and SERCA/PLN), the mechanism of regulation is still unknown. Here, we used a combination of oriented and magic angle spinning (MAS) solid-state NMR spectroscopy to gain atomistic level insights into the mechanism of regulation of SERCA by both PLN and SLN in fully hydrated lipid bilayer membranes.To map protein-protein interactions within lipid membranes, we designed innovative labeling strategies involving various combinations of isotopic labeling schemes, paramagnetic probes, and thioalkylation of SERCA's cysteines. The restraints obtained from the experimental data were used to model the complexes which were further refined using restrained molecular dynamics simulations in explicit lipid bilayers. The structures of SERCA in complex with PLN or SLN provide a detailed view of the binding interface and shed light on the mechanism of SERCA regulation (i.e., inhibition and activation). Knowledge of the biophysical basis of SERCA regulation offers unprecedented opportunity for the rational design of the mutants for gene therapy. We expect the approach and methods used by us to be generally applicable for mapping protein-protein interactions in membrane protein complexes larger than 100 kDa, which are typically inaccessible to other experimental methods.

The Mechanism of Uncoupling ATP Hydrolysis and Calcium Transport in Serca by Sarcolipin

The interactions of sarcolipin (SLN) with the sarco(endo)plasmic reticulum Ca2+ ATPase (SERCA) causes uncoupling of SERCA's ATP hydrolysis and Ca2+ -transport activities. SERCA is an integral component of proper muscle function. It is one of the key proteins responsible for re-establishing the low cytosolic Ca2+ concentrations needed for normal muscle contraction. When SLN is bound to SERCA, fewer moles of Ca2+ are transported per mole of ATP hydrolyzed, which leads to an increase in the heat released. By using solid-state nuclear magnetic resonance spectroscopy and functional assays, we demonstrate that it is the topological coupling between these proteins that is important for the regulation of SERCA. It is through the interactions of their transmembrane regions as well as the interactions of SLN's lumenal tail that lead to uncoupling. These results also provide insight into the mechanism for non-shivering, muscle-based thermogenesis.

Co-Expression of SERCA Isoforms, Phospholamban and Sarcolipin in Human Skeletal Muscle Fibers

Sarcolipin (SLN) and phospholamban (PLN) inhibit the activity of sarco(endo)plasmic reticulum Ca2+-ATPases (SERCAs) by reducing their apparent affinity for Ca2+. A ternary complex between SLN, PLN, and SERCAs results in super-inhibition of SERCA activity. Analysis of skeletal muscle homogenate has limited our current understanding of whether SLN and PLN regulate SERCA1a, SERCA2a, or both in skeletal muscle and whether SLN and PLN are co-expressed in skeletal muscle fibers. Biopsies from human vastus lateralis were analyzed through single fiber Western blotting and immunohisto/fluorescence staining to circumvent this limitation. With a newly generated SLN antibody, we report for the first time that SLN protein is present in human skeletal muscle. Addition of the SLN antibody (50 µg) to vastus lateralis homogenates increased the apparent Ca2+ affinity of SERCA (K Ca, pCa units) (-Ab, 5.85 ± 0.02 vs. +Ab, 5.95 ± 0.02) and maximal SERCA activity (μmol/g protein/min) (-Ab, 122 ± 6.4 vs. +Ab, 159 ± 11) demonstrating a functional interaction between SLN and SERCAs in human vastus lateralis. Specifically, our results suggest that although SLN and PLN may preferentially regulate SERCA1a, and SERCA2a, respectively, physiologically they both may regulate either SERCA isoform. Furthermore, we show that SLN and PLN co-immunoprecipitate in human vastus lateralis homogenate and are simultaneously expressed in 81% of the fibers analyzed with Western blotting which implies that super-inhibition of SERCA may exist in human skeletal muscle. Finally, we demonstrate unequivocally that mouse soleus contains PLN protein suggesting that super-inhibition of SERCA may also be important physiologically in rodent skeletal muscle.

Can proton pumping by SERCA enhance the regulatory role of phospholamban and sarcolipin?

The effect of the incorporation of phosphorylated phospholamban (pPLN) and sarcolipin (SLN) in mercury-supported self-assembled lipid monolayers and in lipid bilayers tethered to mercury via a hydrophilic spacer was investigated by voltammetric techniques and electrochemical impedance spectroscopy. It was shown that pPLN and SLN do not permeabilize lipid bilayers toward ions at physiological pH. However, they exert a permeabilizing action toward inorganic monovalent cations such as K+ and Tl+, but not toward divalent cations such as Ca2+ and Cd2+, following a small decrease in pH. This behavior can be associated with their regulatory action on the Ca-ATPase of the sarcoplasmic reticulum (SERCA). SERCA pumps two Ca2+ ions from the cytosol to the lumen of the sarcoplasmic reticulum (SR) and two protons in the opposite direction, causing a transient decrease of pH in the immediate vicinity of its cytoplasmic domain. This decrease is expected to activate the liberated pPLN molecules and SLN to make the SR membrane leakier toward K+ and Na+ and the SLN ion channel to translocate small inorganic anions, such as Cl−. The effect of pPLN and SLN, which becomes synergic when they are both present in the SR membrane, is expected to favor a rapid equilibration of ions on both sides of the membrane.

New crystal structures of PII-type ATPases: excitement continues

P-type ATPases are ATP-powered ion pumps, classified into five subfamilies (PI-PV). Of these, PII-type ATPases, including Ca2+-ATPase, Na+,K+-ATPase and gastric H+,K+-ATPase, among others, have been the most intensively studied. Best understood structurally and biochemically is Ca2+-ATPase from sarcoplasmic reticulum of fast twitch skeletal muscle (sarco(endo)plasmic reticulum Ca2+-ATPase 1a, SERCA1a). Since publication of the first crystal structure in 2000, it has continuously been a source of excitement, as crystal structures for new reaction intermediates always show large structural changes. Crystal structures now exist for most of the reaction intermediates, almost covering the entire reaction cycle. This year the crystal structure of a missing link, the E1·Mg2+ state, finally appeared, bringing another surprise: bound sarcolipin (SLN). The current status of two other important PII-type ATPases, Na+,K+-ATPase and H+,K+-ATPase, is also briefly described.

Sarcolipin trumps β‐adrenergic receptor signaling as the favored mechanism for muscle‐based diet‐induced thermogenesis

Sarcolipin (SLN) regulates muscle-based nonshivering thermogenesis and is up-regulated with high-fat feeding (HFF). To investigate whether other muscle-based thermogenic systems compensate for a lack of Sln and to firmly establish SLN as a mediator of diet-induced thermogenesis (DIT), we measured muscle and whole-body energy expenditure in chow- and high-fat-fed Sln(-/-) and wild-type (WT) mice. Following HFF, resting muscle metabolic rate (VO2, μl/g/s) was increased similarly in WT (0.28±0.02 vs. 0.31±0.03) and Sln(-/-) (0.23±0.03 vs. 0.35±0.02) mice due to increased sympathetic nervous system activation in Sln(-/-) mice; however, whole-body metabolic rate (VO2, ml/kg/h) was lower in Sln(-/-) compared with WT mice following HFF but only during periods when the mice were active in their cages (WT, 2894±87 vs. Sln(-/-), 2708±61). Treatment with the β-adrenergic receptor (β-AR) antagonist propranolol during HFF completely prevented muscle-based DIT in Sln(-/-) mice; however, it had no effect in WT mice, resulting in greater differences in whole-body metabolic rate and diet-induced weight gain. Our results suggest that β-AR signaling partially compensates for a lack of SLN to activate muscle-based DIT, but SLN is the primary and more effective mediator.

Ablation of sarcolipin decreases the energy requirements for Ca2+ transport by sarco(endo)plasmic reticulum Ca2+-ATPases in resting skeletal muscle

The purpose of this study was to examine the effects of sarcolipin (SLN) on sarco(endo) plasmic reticulum Ca2+-ATPase (SERCA pump) energetics in vivo and resting skeletal muscle metabolic rate. Using SLN knockout (Sln−/−) mice we show that SLN ablation increases the transport stoichiometry of SERCA pumps (Ca2+ uptake/Ca2+-ATPase activity) and decreases the relative contribution of SERCA pumps to resting oxygen consumption (VO2) in soleus without affecting soleus or whole body VO2. These data suggest that the lower energy requirements for Ca2+ cycling in resting skeletal muscle of Sln−/− mice do not impact significantly either skeletal muscle or whole body metabolic rate.