Journal of Biological Chemistry
Volume 277, Issue 47, 22 November 2002, Pages 44740-44746
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MEMBRANE TRANSPORT STRUCTURE FUNCTION AND BIOGENESIS
Sarcolipin Overexpression in Rat Slow Twitch Muscle Inhibits Sarcoplasmic Reticulum Ca2+ Uptake and Impairs Contractile Function*

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Sarcolipin (SLN) is an inhibitor of sarco(endo)plasmic reticulum Ca2+-ATPases (SERCAs)in vitro, but its function in vivo has not been defined. NF-SLN cDNA (SLN tagged N-terminally with a FLAG epitope) was introduced into rat soleus muscle in one hindlimb by plasmid injection and electrotransfer. Western blotting showed expression and co-immunoprecipitation showed physical interaction between NF-SLN and SERCA2a. Contractile properties and SERCA2a function were assessed and compared with vector-injected contralateral soleus muscles. NF-SLN reduced both peak twitch force (P t) (123.9 ± 12.5 versus 69.8 ± 8.9 millinewtons) and tetanic force (P o) (562.3 ± 51.0versus 300.7 ± 56.9 millinewtons) and reduced both twitch and tetanic rates of contraction (+dF/dt) and relaxation (−dF/dt) significantly. Repetitive stimulation (750-ms trains at 50 Hz once every 2 s for 3 min) showed that NF-SLN increased susceptibility to fatigue. These changes in contractile function were observed in the absence of endogenous phospholamban, and NF-SLN had no effect on either SERCA2a or SERCA1a expression levels. NF-SLN also decreased maximal Ca2+transport activity at pCa 5 by 31% with no significant change in apparent Ca2+ affinity (6.36 ± 0.07versus 6.39 ± 0.08 pCa units). These results show that NF-SLN expression impairs muscle contractile function by inhibiting SERCA function and diminishing sarcoplasmic reticulum Ca2+ stores.

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*

This work was supported by Grant MT12545 from the Canadian Institutes of Health Research and Grant T5042 from the Heart and Stroke Foundation of Ontario (to D. H. M.).

These authors contributed equally to this work.

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Postdoctoral Fellow of the Heart and Stroke Foundation of Canada.