Résumé :
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Muscle contraction relies on efficient calcium release from sarcoplasmic reticulum (SR), performed by the ryanodine receptor (RyR1) upon cell stimulation. RyR1 is 5000 amino acids long, anchored in the membrane of SR, and coupled to the voltage-sensitive calcium channel DHPR that initiate muscle contraction after depolarisation of the fibre. Several mutations in the RyR1 gene have been associated to inherited muscle defects, mostly congenital structural myopathies (central core disease, CCD, and multi minicore disease, MmD) and malignant hyperthermia. Although mutations are found along the whole RYR1 gene, those associated with CCD and MmD seem to cluster in the 3’ region coding for transmembrane domains of the protein. To date, most of the studies on mutations in these regions have led to the hypothesis that mutated RyR1 is defective in its channel properties. The 14646+2.99 kb A->G mutation has been found in a patient with severe MmD. It generates a cryptic splicing site that leads to an out-of-frame insertion of one exon in the most 3’ end of the RyR1 cDNA. The expected protein is shorter than normal RyR1 and lacks its last transmembrane helix (TM6). Moreover, in a biopsy of this patient, the overall level of RyR1 is drastically reduced. To understand this reduction in RyR1 level, we have tested whether removal of TM6 helix could destabilise the mutated RyR1 (mRyR). Surprisingly, when expressed in a cell line, the mRyR construct is mislocalised, compared to normal RyR1 present in the endoplasmic reticulum. We have investigated the precise intracellular targeting of mRyR1 as well as the mechanisms leading to its accumulation. Our work raises the question whether, in addition to channel defaults, intracellular localisation of mutated RyR1 could also explain some of the pathological effects for mutations associated with congenital myopathies.
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