Characterization of the 70-kDa peroxisomal membrane protein, an ATP binding cassette transporter

The 70-kDa peroxisomal membrane protein (PMP70) is one of the major compone nts of rat liver peroxisomal membranes and belongs to a superfamily of prot eins known as ATP binding cassette transporters. PMP70 is markedly induced by administration of hypolipidemic agents in parallel with peroxisome proli feration and induction of peroxisomal fatty acid beta-oxidation enzymes, To characterize the role of PMP70 in biogenesis and function of peroxisomes, we transfected the cDNA of rat PMP70 into Chinese hamster ovary cells and e stablished cell lines stably expressing PMP70. The content of PMP70 in the transfectants increased about 5-fold when compared with the control cells. A subcellular fractionation study showed that overexpressed PMP70 was enric hed in peroxisomes, This peroxisomal localization was confirmed by immunofl uorescence and immunoelectron microscopy. The number of immune-gold particl es corresponding to PMP70 on peroxisomes increased markedly in the transfec tants, but the size and the number of peroxisomes were essentially the same in both the transfectants and the control cells. beta-Oxidation of palmiti c acid increased about 2-3-fold in the transfectants, whereas the oxidation of lignoceric acid decreased about 30-40%. When intact peroxisomes prepare d from both the cell lines were incubated with palmitoyl-CoA, oxidation was stimulated with ATP, but the degree of the stimulation was higher in the t ransfectants than in the control cells. Furthermore, we established three C hinese hamster ovary cell lines stably expressing mutant PMP70. In these ce lls, beta-oxidation of palmitic acid decreased markedly, These results sugg est that PMP70 is involved in metabolic transport of long chain acyl-CoA ac ross peroxisomal membranes and that increase of PMP70 is not associated wit h proliferation of peroxisomes.