We are studying the chronic regulation of mitochondrial enzyme activity in muscle, with a view to understanding the decreased capacity of skeletal muscle to respond to endurance training in old age.
We aim to identify signals which allow the cell to sense increased energy demand and respond with increased expression of mitochondrial proteins encoded by the nuclear and mitochondrial genomes. This year we have studied C2C12 cells in culture (a mouse muscle-derived cell line) and have shown that differentiation from a tumor-like to a myocyte-like morphology, in response to serum-starvation, precedes the induction of mitochondrial enzyme activity. Similarly, in NIH 3T3 cells, which we have investigated because they show a very large (up to 40-fold) induction of mitochondrial enzymes upon differentiation into an adipocyte-like phenotype, the induction of lipogenic capacity precedes the mitochondrial changes. Thus, it is likely that the energy-demand of differentiation induces the mitochondrial enzyme synthesis. We established that inner mitochondrial membrane enzymes with subunits coded for on mt-DNA (Complexes I and IV) show a lag in synthesis during differentiation in both cell types, whereas nuclear encoded matrix enzymes (citrate synthase, NAD-isocitrate dehydrogenase, pyruvate dehydrogenase, 3-hydroxyacyl CoA dehydrogenase) increase in a linear way. Succinate dehydrogenase (Complex II), which is a membrane enzyme with no mt-DNA-encoded subunit, behaves like the matrix enzymes, suggesting that it is the distinction between mt-DNA and nuclear DNA which is important, rather than the localization of the protein. Mitochondrial membrane enzyme activities are still much lower in differentiated C2C12 cells than in mouse soleus muscle, raising the question of an additional role of membrane lipid in organizing functional enzyme complexes - which we are investigating. Northern blots for COX II - a subunit encoded on mt-DNA - shows that this message tracks the copy number of mt-DNA in differentiating C2C12 cells, suggesting a rather straight-forward mode of control, at least in this instance. Northern blots for other representative enzymes are in progress.
