Evidence that phosphatidylserine is imported into mitochondria via a mitochondria-associated membrane and that the majority of mitochondrial phosphatidylethanolamine is derived from decarboxylation of phosphatidylserine.

Phosphatidylserine is synthesized in both the endoplasmic reticulum and a unique membrane fraction, the mitochondria-associated membrane (MAM) (Vance, J.E. (1990) J. Biol. Chem. 265, 7248-7256). In Chinese hamster ovary cells labeled with [3H]serine or [3H]ethanolamine, we found that the majority of mitochondrial phosphatidylethanolamine was derived from phosphatidylserine decarboxylation. Essentially no mitochondrial phosphatidylethanolamine, especially that in the inner membrane, was imported from the endoplasmic reticulum. We tested the hypothesis that phosphatidylserine made in the endoplasmic reticulum is delivered via the MAM to mitochondria for decarboxylation to phosphatidylethanolamine. Cells were pulse-labeled with [3H]serine and subsequently incubated either in the presence of hydroxylamine (for inhibition of phosphatidylserine decarboxylation) or under conditions for which cellular ATP had been depleted (for inhibition of phosphatidylserine import into mitochondria). In hydroxylamine-treated cells, within 2 h, the amount of radiolabeled phosphatidylserine in the MAM and mitochondria, but not microsomes, was greater than in untreated cells. Moreover, in ATP-depleted, but not in control, cells the amount of radiolabeled phosphatidylserine in the MAM approximately doubled by 3 h. These observations are consistent with the hypothesis that newly synthesized phosphatidylserine normally traverses the MAM en route to mitochondria.