Enhanced Ca²⁺ transport and muscle relaxation in skeletal muscle from sarcolipin-null mice

Abstract

To evaluate the physiological significance of SLN in skeletal muscle, we compared muscle contractility and SERCA activity between Sln-null and wild-type mice. SLN protein expression in wild-type mice was abundant in soleus and red gastrocnemius (RG), low in extensor digitorum longus (EDL), and absent from white gastrocnemius (WG). SERCA activity rates were increased in soleus and RG, but not in EDL or WG, from Sln-null muscles, compared with wild type. No differences were seen between wild-type and Sln-null EDL muscles in force-frequency curves or maximum rates of force development (ϩdF/dt). Maximum relaxation rates (ϪdF/dt) of EDL were higher in Sln-null than wild type across a range of submaximal stimulation frequencies, but not during a twitch or peak tetanic contraction. For soleus, no differences were seen between wild type and Sln-null in peak tetanic force or ϩdF/dt; however, forcefrequency curves showed that peak force during a twitch and 10-Hz contraction was lower in Sln-null. Changes in the soleus forcefrequency curve corresponded with faster rates of force relaxation at nearly all stimulation frequencies in Sln-null compared with wild type. Repeated tetanic stimulation of soleus caused increased (ϪdF/dt) in wild type, but not in Sln-null. No compensatory responses were detected in analysis of other Ca 2ϩ regulatory proteins using Western blotting and immunohistochemistry or myosin heavy chain expression using immunofluorescence. These results show that 1) SLN regulates Ca 2ϩ -ATPase activity thereby regulating contractile kinetics in at least some skeletal muscles, 2) the functional significance of SLN is graded to the endogenous SLN expression level, and 3) SLN inhibitory effects on SERCA function are relieved in response to repeated contractions thus enhancing relaxation rates. knockout mouse; muscle contractility; isolated skeletal muscle; Ca 2ϩ pump SARCO (ENDO) PLASMIC RETICULUM (SER) Ca 2ϩ -ATPases (SERCAs) are ubiquitously expressed, integral membrane proteins that transport Ca 2ϩ ions from the cytosol to the lumen of the SER. In the heart, a 52 amino acid transmembrane protein, phospholamban (PLN), interacts physically with SERCA2a, lowering the apparent Ca 2ϩ affinity of the PLN-SERCA2a complex (19,30). The inhibited complex is disrupted by phosphorylation of PLN or by elevation of cytosolic Ca 2ϩ , leading to the reversal of SERCA2a inhibition. Sarcolipin (SLN), a 31 amino acid protein, shares substantial identity with PLN in both primary sequence and gene structure (25, 35) and, like PLN, is an effective inhibitor of SERCA molecules (1-3, 16, 24). SLN was originally identified as a proteolipid that copurified with SERCA1a in rabbit fast-twitch skeletal muscle (20). Subsequently, SLN was found to be expressed highly in both human and rabbit fast-twitch skeletal muscle and to a lesser extent in slow-twitch and cardiac muscle, based on mRNA levels (25). SLN expression in the heart is largely restricted to the atrium (22), whereas PLN is highly expressed in the ventricle and not in the atrium ...

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