Transbilayer movement of phosphatidylcholine in the mitochondrial outer membrane of Saccharomyces cerevisiae is rapid and bidirectional

The process of transmembrane movement of phosphatidylcholine (PC) across th e outer membrane of mitochondria was investigated in vitro in mitochondrial outer membrane vesicles (OMV) from the yeast Saccharomyces cerevisiae. Pho sphatidylcholine-transfer protein (PC-TP) was used to extract radiolabeled PC from OMV, with small unilamellar vesicles serving as acceptor system. En dogenously radiolabeled PC synthesized either via the CDP-choline pathway o r via methylation of phosphatidylethanolamine can be extracted completely f rom the OMV with a t(1/2) of 1 min or less at 30 degrees C. The size of the pool of PC in OMV available for exchange by PC-TP is not affected by pretr eatment of the OMV with proteinase It or sulfhydryl reagents. In the revers e experiment where radiolabeled PC was introduced into the OMV, similar cha racteristics for the exchange were found. The accessibility of labeled PC t o externally added phospholipase A(2) was used as a measure for its transme mbrane distribution. It was found that PC is not exclusively located in the outer leaflet of the OMV. Only 30-35% can be degraded in intact OMV by pho spholipase A(2), irrespective of whether the PC is introduced by PC-TP or e ndogenously synthesized via either of the pathways of biosynthesis. The res ults demonstrate the occurrence of rapid bidirectional transbilayer movemen t of both endogenous and in vitro introduced PC in OMV. Furthermore, there appears to be no preference for mitochondrial import of PC synthesized by e ither of the pathways in vivo. (C) 1999 Elsevier Science B.V. All rights re served.