Molecular cloning of the rat NADH:ubiquinone oxidoreductase subunit and its up-regulation in the facial muscle after denervation: detected by means of differential display.

The differential display method was applied to identify gene expression which is especially up-regulated in the rat denervated skeletal muscle. Total RNA from normal and denervated facial muscles was isolated, amplified by PCR using certain primers, and separated by electrophoresis on polyacrylamide gel. PCR products which were apparently higher in the operated than in the control side were cut out from the gel and subsequently sequenced. One of the cDNA fragments obtained in the present study showed about 80% identity in nucleotide sequence and about 84% identity in amino acid sequence to one of the NADH: ubiquinone oxidoreductase (complex I) subunits from the bovine heart mitochondria (813). Complex I, the first and largest enzyme of the mitochondrial respiratory chain, transfers electrons from NADH to ubiquinone-10 (UQ-10). B13 is a 13 kDa subunit; deficiency of which causes incomplete or incorrect assembly of a functional complex I, resulting in one of the types of human mitochondrial myopathy. Northern blot analysis and in situ hybridization histochemistry confirmed that rat B13 mRNA expression was up-regulated after denervation and was distributed throughout whole muscle cell body. These findings suggest that up-regulation of the rat B13 mRNA expression after denervation plays an important role in the effective electron transfer from NADH to UQ-10 in skeletal muscle cells, resulting in restriction of lipid peroxidation and biological tissue damage.