Résumé :
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Communication n° 415. The sarcoplasmic reticulum (SR) contains major proteins that control the intracellular Ca2+ concentration and play an important role in excitation-contraction coupling mechanism (ECC). Several less abundant components have been identified, but their role still remains unclear. However, because of their localization they could be involved in ECC and regulation of Ca2+ homeostasis; thus a deficiency in their expression or function could be associated with neuromuscular diseases. Junctate is a newly identified integral SR/ER membrane calcium binding protein, which is an alternative splicing form of the same gene generating aspartyl-b-hydroxylase and junctin. It is expressed in a variety of tissues and at low levels in skeletal muscle. The aim of this project is to study the functional role of junctate in the mechanisms of regulation of intracellular Ca2+ concentration in skeletal muscle, using a transgenic mouse model characterized by an over-expression of junctate. Several lines of transgenic mice were generated using junctate cDNA under the control of a skeletal muscle specific promoter. The insertion of the transgene in genomic DNA was detected by PCR and homozygous mice identified by southern blot. The validity of this animal model was confirmed by western blot analysis which supported over-expression of junctate in skeletal muscle. Study of Ca2+ loading and release processes performed on SR vesicles, using a spectrophotometric determination with Antipyrylazo-III, showed that maximal Ca2+ loading was significantly increased in transgenic mice compared to their wild type counterpart. In addition, the data indicated a decrease in the extent of Ca2+ release induced by 4-chloro-m-cresol in muscle preparation from transgenic mice. These findings demonstrate that over-expression of junctate in skeletal muscle modifies the Ca2+ loading characteristics and the total quantity of Ca2+ release, indicating that junctate might have a role in Ca2+ homeostasis in skeletal muscle.
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