FLUORESCENTLY LABELED PHOSPHATIDYLINOSITOL TRANSFER PROTEIN (ISOFORM-ALPHA AND ISOFORM-BETA), MICROINJECTED INTO FETAL BOVINE HEART ENDOTHELIAL-CELLS, ARE TARGETED TO DISTINCT INTRACELLULAR SITES

Upon permeabilization of Swiss mouse 3T3 fibroblasts, an isoform of ph osphatidylinositol transfer protein (PI-TP) was preferentially retaine d, a major part of which was associated with the perinuclear Golgi sys tem (K. J. de Vries, A. Momchilova-Pankova, G. T. Snoek, and K. W. A. Wirtz, Exp. Cell Res. 215, 109-113, 1994). In the present study, the i ntracellular localization of this isoform (PI-TP beta) and the regular form (PI-TP alpha) was investigated in fetal bovine heart endothelial cells by microinjection of fluorescently labeled analogs followed by confocal laser scanning microscopy. The PI-TP alpha and PI-TP beta use d were purified from bovine brain cytosol and covalently labeled with sulfoindocyanine dyes. By this novel method it was found that PI-TP be ta was preferentially associated with perinuclear membrane structures whereas PI-TP alpha was predominantly present in the nucleus and in th e cytoplasm. This intracellular localization was confirmed by indirect immunofluorescence indicating that the fluorescently labeled PI-TP al pha and PI-TP beta were targeted to the same sites as their endogeneou s counterparts. (C) 1996 Academic Press, Inc.