TRANSFECTION OF THE KETOGENIC MITOCHONDRIAL 3-HYDROXY-3-METHYLGLUTARYL-COENZYME-A SYNTHASE CDNA INTO MEV-1 CELLS CORRECTS THEIR AUXOTROPHY FOR MEVALONATE

A somatic cell mutant of the Chinese hamster ovary (CHO)-K1 (called Me v-1), auxotrophic for mevalonate by virtue of a complete lack of detec table cytosolic 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) syntha se activity, was transfected with a plasmid containing the cDNA for ke togenic mitochondrial HMG-CoA synthase under the control of SV40 early promoter. The resulting stable cell line (Mev-SM) was able to grow in the absence of mevalonate. Analysis by Western blot showed that the n ew cell line strongly expressed mitochondrial HMG-CoA synthase protein . Immunocytochemical studies using specific antibodies against mitocho ndrial HMG-CoA synthase showed that the protein was located exclusivel y inside the mitochondria. The prototroph cell line Mev-SM can incorpo rate labeled ac etate into cholesterol in the absence of mevalonate. T hese results show that the new cell, line may circumvent the lack of c ytosolic HMG-CoA synthase activity by producing cholesterol-convertibl e HMG-CoA inside the mitochondria.