Beręsewicz Małgorzata, Boratyńska-Jasińska Anna, Charzewski Łukasz, Kawalec Maria, Kabzińska Dagmara, Kochański Andrzej, Krzyśko Krystiana A, Zabłocka Barbara
Molecular Biology Unit, Mossakowski Medical Research Centre, PAS, Warsaw, Poland.
Department of Physics, University of Warsaw, Warsaw, Poland.
PLoS One. 2017 Jan 11;12(1):e0169999. doi: 10.1371/journal.pone.0169999. eCollection 2017.
Charcot-Marie-Tooth disease type 2A (CMT2A) is an autosomal dominant axonal peripheral neuropathy caused by mutations in the mitofusin 2 gene (MFN2). Mitofusin 2 is a GTPase protein present in the outer mitochondrial membrane and responsible for regulation of mitochondrial network architecture via the fusion of mitochondria. As that fusion process is known to be strongly dependent on the GTPase activity of mitofusin 2, it is postulated that the MFN2 mutation within the GTPase domain may lead to impaired GTPase activity, and in turn to mitochondrial dysfunction. The work described here has therefore sought to verify the effects of MFN2 mutation within its GTPase domain on mitochondrial and endoplasmic reticulum morphology, as well as the mtDNA content in a cultured primary fibroblast obtained from a CMT2A patient harboring a de novo Arg274Trp mutation. In fact, all the parameters studied were affected significantly by the presence of the mutant MFN2 protein. However, using the stable model for mitofusin 2 obtained by us, we were next able to determine that the Arg274Trp mutation does not impact directly upon GTP binding. Such results were also confirmed for GTP-hydrolysis activity of MFN2 protein in patient fibroblast. We therefore suggest that the biological malfunctions observable with the disease are not consequences of impaired GTPase activity, but rather reflect an impaired contribution of the GTPase domain to other MFN2 activities involving that region, for example protein-protein interactions.
2A型夏科-马里-图斯病（CMT2A）是一种常染色体显性遗传性轴索性周围神经病，由线粒体融合蛋白2基因（MFN2）突变引起。线粒体融合蛋白2是一种存在于线粒体外膜的GTP酶蛋白，通过线粒体融合负责调节线粒体网络结构。由于已知该融合过程强烈依赖于线粒体融合蛋白2的GTP酶活性，因此推测GTP酶结构域内的MFN2突变可能导致GTP酶活性受损，进而导致线粒体功能障碍。因此，本文所述工作旨在验证GTP酶结构域内的MFN2突变对线粒体和内质网形态以及从一名携带新生Arg274Trp突变的CMT2A患者获得的原代培养成纤维细胞中线粒体DNA含量的影响。事实上，所有研究参数均受到突变型MFN2蛋白的显著影响。然而，利用我们获得的线粒体融合蛋白2稳定模型，我们接下来能够确定Arg274Trp突变不会直接影响GTP结合。患者成纤维细胞中MFN2蛋白的GTP水解活性也证实了这些结果。因此，我们认为该疾病中可观察到的生物学功能障碍不是GTP酶活性受损的后果，而是反映了GTP酶结构域对涉及该区域的其他MFN2活性（例如蛋白质-蛋白质相互作用）的贡献受损。
