Characterization of a Chinese Hamster Mutant in Mitochondrial Gene Expression

Abstract

Gal\u2732 is a Chinese hamster lung (CHL} cell mutant that is unable to grow in galactose due to a defect in mitochondria! protein synthesis, In this unique CHL mutant, the mitochondrially synthesized subunits of NADH dehydrogenase and cytochrome c oxidase are significantly reduced when compared to the wild-type levels, but not the ATPase subunits. In contrast to the protein levels, the mRNA levels as determined by Northern and slot blotting are elevated in the mutant compared to the wild type, Thus, the decreased levels of mitochondrially synthesized proteins in Gal 32 are the result of decreased translation of elevated mRNAs. To determine whether the differentially reduced protein synthesis in Gal\u2732 is at the level of translational initiation or elongation or sequestering of mRNA, the polysome profile of the Gal 32 mutant is compared to that of wild type. The polysome profile which is generated by centrifugation through a 1 5-30% sucrose gradient is analyzed by Northern blotting using cytochrome oxidase, NADH dehydrogenase, ATPase riboprobes and 16S and 12S DNA probes. The location and amount of the mRNAs and rRNAs are being examined to understand the molecular mechanism of decreased translation. Supported by NIH-MBRS S06-GM08016