La dystrophie musculaire d'Emery-Dreifuss (EDMD) est caractérisée par des retractions précoces, une faiblesse et atrophie musculaire lentement progressive, et une atteinte cardiaque. Les mutations des gènes EMD et LMNA sont respectivement responsables de formes liées à l'X et de formes autosomiques de l'EDMD. Ces deux gènes codent pour des protéines de l'enveloppe nucléaire, l'émerine et les lamines A/C. Les mutations du gène FHL1 ont été impliquées dans d'autres cas d'EDMD liée à l'X. FHL1 codent pour FHL1A, FHL1B et FHL1C, protéines jamais décrites comme localisées à l'enveloppe nucléaire. Nous avons cherché à enrichir les connaissances sur la distribution subcellulaire des différentes isoformes de FHL1 dans les muscles squelettiques humains sains et malades. Nous avons mis en évidence que les isoformes FHL1 présentent à la fois une localisation cytoplasmique et nucléaire dans les myoblastes humains. Au noyau, FHL1B est fortement accumulé au niveau de l'enveloppe nucléaire où il interagit avec les lamines A/C et l'émerine. Cette localisation à l'enveloppe nucléaire est indépendante de l'expression de l'émerine ou des lamines A/C. La différenciation des myoblastes entraîne une forte réduction de l'expression de FHL1B et de son exclusion progressive du noyau, n'impliquant pas la protéine CRM-1. Nous avons mis en évidence l'augmentation de l'expression de FHL1B dans les myoblastes de deux patients atteints d'EDMD, l'un porteur d'une mutation dans le gène LMNA, l'autre dans le gène FHL1. En conclusion, la localisation spécifique de FHL1B et sa modulation dans les myoblastes de patients confirment les cas d'EDMD liés à FHL1 comme des pathologies de l'enveloppe nucléaire. / Emery-Dreifuss muscular dystrophy (EDMD) is characterized by the triad of early contractures, slowly progressive muscle wasting and weakness, and cardiac disease. Mutations in EMD and LMNA, encoding for the nuclear envelope (NE) proteins emerin and lamin A/C, are associated with X-linked and autosomal form of EDMD, respectively. The discovery that FHL, encoding FHL1A, FHL1B and FHL1C, is implicated in the pathogenesis of EDMD, raises the question of how a non-NE protein can be linked to emerin and lamin A/C. We aimed to provide knowledge of the subcellular distribution and expression of the various FHL1 isoforms in healthy and diseased human skeletal muscle. We found that FHL1 isoforms display a dual cytoplasmic and nuclear localization in human myoblasts. In addition, FHL1B strongly accumulated at the NE where it interacted with both lamin A/C and emerin. NE localization of FHL1B was independent of emerin and lamin A/C expression. Myoblast differentiation resulted in greatly reduced FHL1B protein expression and in the progressive nuclear exclusion of FHL1 protein isoforms. We have shown that chromosome region maintenance 1 (CRM1)-mediated nuclear export was not involved in the progressive decrease of nucleoplasmic FHL1B. Finally, we detected increased FHL1B protein levels in myoblasts of two patients with LMNA- and FHL1-related EDMD. Altogether, the specific localization of FHL1B and its modulation in disease-patient’s myoblasts confirmed FHL1-related EDMD as a NE disease.

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It is known that muscles that lack KATP channel activity generate much greater unstimulated [Ca2+]i and force than normal muscles during fatigue. The increase in unstimulated force in KATP channel deficient muscles is abolished by a partial inhibition of L-type Ca2+ channels, suggesting that it is due to a Ca2+ influx through L-type Ca2+ channels and a subsequent increased myoplasmic Ca2+. However, there is also evidence that the increase in resting force is abolished by NAC, a ROS scavenger. The objective of this study was to reconcile these observations by studying the hypothesis that “the increase in resting [Ca2+]i during fatigue in KATP channel deficient muscles starts with an excess Ca2+ influx through L-type Ca2+ channels, followed by an excess ROS production that causes a further increase in resting [Ca2+]i”. To test the hypothesis, single FDB fibres were fatigued with one tetanic contraction/sec for 180 sec. KATP channel deficient fibres were obtained i) by exposing wild type muscle fibers to glibenclamide, a KATP channel blocker and ii) by using fibres from Kir6.2-/- mice, which are null mice for the Kir6.2 gene that encodes for the protein forming the channel pore. Verapamil, a L-type Ca2+ channel blocker, applied at 1 μM, significantly reduced resting [Ca2+]i during fatigue in glibenclamide-exposed wild type fibres. NAC (1 mM) also reduced resting [Ca2+]i in glibenclamide-exposed muscles. The results suggest that the increase in resting [Ca2+]i during fatigue in KATP channel deficient FDB fibres is due to an influx through L-type Ca2+ channels, and an excess ROS production.

It is known that muscles that lack KATP channel activity generate much greater unstimulated [Ca2+]i and force than normal muscles during fatigue. The increase in unstimulated force in KATP channel deficient muscles is abolished by a partial inhibition of L-type Ca2+ channels, suggesting that it is due to a Ca2+ influx through L-type Ca2+ channels and a subsequent increased myoplasmic Ca2+. However, there is also evidence that the increase in resting force is abolished by NAC, a ROS scavenger. The objective of this study was to reconcile these observations by studying the hypothesis that “the increase in resting [Ca2+]i during fatigue in KATP channel deficient muscles starts with an excess Ca2+ influx through L-type Ca2+ channels, followed by an excess ROS production that causes a further increase in resting [Ca2+]i”. To test the hypothesis, single FDB fibres were fatigued with one tetanic contraction/sec for 180 sec. KATP channel deficient fibres were obtained i) by exposing wild type muscle fibers to glibenclamide, a KATP channel blocker and ii) by using fibres from Kir6.2-/- mice, which are null mice for the Kir6.2 gene that encodes for the protein forming the channel pore. Verapamil, a L-type Ca2+ channel blocker, applied at 1 μM, significantly reduced resting [Ca2+]i during fatigue in glibenclamide-exposed wild type fibres. NAC (1 mM) also reduced resting [Ca2+]i in glibenclamide-exposed muscles. The results suggest that the increase in resting [Ca2+]i during fatigue in KATP channel deficient FDB fibres is due to an influx through L-type Ca2+ channels, and an excess ROS production.