Phosphatidylinositol transfer protein in murine embryonal carcinoma cells during retinoic acid-induced differentiation |
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Authors: | I. Suzanna C. de Wit Siegfried W. de Laat Gerry T. Snoek Karel W. A. Wirtz |
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Affiliation: | Centre for Biomembranes and Lipid Enzymology, Utrecht University, Padualaan 8, 3584 CT Utrecht, The Netherlands.;Hubrecht Laboratory, Netherlands Institute for Developmental Biology, Uppsalalaan 8, 3584 CT Utrecht, The Netherlands. |
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Abstract: | Phosphatidylinositol transfer protein (PI-TP) was studied in P19 embryonal carcinoma (EC) cells at different stages of retinoic acid (RA) induced differentiation. Western blot analysis indicated an increased expression of PI-TP (35 kDa) during differentiation. Western blots of isoelectric focusing gels showed that the 35 kDa band consisted of the PI-carrying form of PI-TP (pl 5.5) and of a novel, more acidic form of PI-TP (pl 5.4), levels of both of which increased during differentiation. These increased levels were not reflected in the in vitro PI-transfer activity of the cytosolic fraction nor in the mRNA levels as analyzed by northern blotting. By using indirect immunofluorescence it was shown that PI-TP is localized in the cytoplasm and associated with perinuclear Golgi structures and that this distribution is slightly affected during RA-induced differentiation. Immunoprecipitation of PI-TP from [32P]Pi labeled cells demonstrated that the level of phosphorylation of PI-TP is high in undifferentiated P19 EC cells and low after 5 days of RA-induced differentiation. These results strongly suggest that changes in the levels of PI-TP are intimately connected with changes in the growth characteristics of P19 EC cells during RA-induced differentiation. It remains to be established to what extent this connection is governed by the recent finding that PI-TP is an essential cytosolic factor in stimulating phospholipase C activity. |
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Keywords: | immunofluorescence P19 EC cells phosphatidylinositol transfer protein RA-induced differentiation |
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