The construction and preliminary investigation of the cell model of a novel mutation R675Q in the SCN4A gene identified in a Chinese family with normokalemic periodic paralysis / 中华医学遗传学杂志

ABSTRACT

<p><b>OBJECTIVE</b>To construct and investigate the cell model of a novel mutation R675Q in the skeletal muscle Na channel type 4 alpha subunit gene (SCN4A) identified from a Chinese family with normokalemic periodic paralysis.</p><p><b>METHODS</b>cDNA encoding the adult isoform of SCN4A was used as a template for in vitro site-directed mutagenesis by PCR method. The mutated plasmid was transiently transfected into HEK-293 cells by calcium phosphate precipitation. Twenty four and 48 hours after transfection, the expression level of SCN4A was detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. Whole cell voltage-clamp recording was used to study the current of sodium channels.</p><p><b>RESULTS</b>The site-mutagenesis of the plasmid was confirmed by sequencing. The expression of SCN4A gene was significantly elevated 24 h and 48 h after transfection. The relative current of R675Q is smaller than that of wide type before reaching peak current under the same test voltage, but larger than that of wild type current after reaching peak current. They both had the largest peak current under 0 mV test pulse.</p><p><b>CONCLUSION</b>A cell model of normokalemic periodic paralysis was successfully constructed. The R675Q mutation of the SCN4A gene enhances the activation and inactivation of the sodium channel, and the S4 transmembrane segment may have intimate relationship with the attack of weakness in normoKPP patients.</p>