Protein Import into Subsarcolemmal and Intermyofibrillar Skeletal Muscle Mitochondria: DIFFERENTIAL IMPORT REGULATION IN DISTINCT SUBCELLULAR REGIONS

Abstract

To date, no studies have described the import of proteins in mitochondria obtained from skeletal muscle. In this tissue, mitochondria consist of the functionally and biochemically distinct intermyofibrillar (IMF) and subsarcolemmal (SS) subfractions, which are localized in specialized cellular compartments. This mitochondrial heterogeneity in muscle could be due, in part, to differential rates of protein import. To evaluate this possibility, the import of precursor malate dehydrogenase and ornithine carbamyltransferase proteins was investigated in isolated IMF and SS mitochondria in vitro. Import of these was 3-4-fold greater in IMF compared with SS mitochondria as a function of time. This could account for the higher malate dehydrogenase enzyme activity in IMF mitochondria. Divergent import rates in IMF and SS mitochondria likely result from a differential reliance on various components of the import pathway. SS mitochondria possess a greater content of the molecular chaperones hsp60 and Grp75, yet import is lower than in IMF mitochondria. On the other hand, adriamycin inhibition studies illustrated a greater reliance on acidic phospholipids (i.e. cardiolipin) for the import process in SS mitochondria. Matrix ATP levels were 3-fold higher in IMF mitochondria, but experiments in which ATP depletion was performed with atractyloside and oligomycin illustrated a dissociation between import rates and levels of ATP. In contrast, a close relationship was found between the rate of ATP production (i.e. mitochondrial respiration) and protein import. When respiratory rates in IMF and SS mitochondria were equalized, import rates in both subfractions were similar. These data indicate that 1) import rates are more closely related to the rate of ATP production than the steady state ATP level, 2) import into IMF and SS mitochondrial subfractions is regulated differently, and 3) mitochondrial heterogeneity within a cell type can be due to differences in the rates of protein import, suggesting that this step is a potentially regulatable event in determining the final mitochondrial phenotype.