INFLUENCE OF SUBUNIT TRANSCRIPT AND PROTEIN-LEVELS ON FORMATION OF A MITOCHONDRIAL MULTIENZYME COMPLEX

Constitutive expression of nuclear genes encoding mitochondrial protei ns raises the question of whether these proteins are present in simila r amounts in mitochondria of different tissues. We report that amounts of a single multienzyme complex can vary on a per mitochondrion basis depending on the number of mitochondria per cell. Human branched-chai n alpha-keto acid dehydrogenase (BCKD) expression is used as a paradig m in these studies. Expression is compared and contrasted in HepG2 and DG75 cells in which mitochondrial content is twofold higher in the he patocarcinoma line than in the lymphoblastoid line. Per cell, BCKD act ivity is equal in the two cell types, but BCKD protein concentration p er mitochondrion is twofold higher in DG75 cells. Steady-state mRNA le vels do not appear to be directly related to amounts of protein in the two cell lines. To test whether one subunit is limiting in formation of complex, overexpression of each BCKD subunit was elicited by plasmi d transfection of the DG75 cells. Only overexpression of the beta-subu nit of the decarboxylase component induced more BCKD activity without apparent increase in mRNA for the other endogenously expressed subunit s. This implies that free BCKD subunits exist in a cell and can be rec ruited into an active complex when the limiting subunit becomes availa ble. (C) 1996 Wiley-Liss, Inc.