Expression of glycoconjugates bearing the Lewis X epitope during neural differentiation of P19 EC cells

Abstract

The Lewis X (Le x) bearing glycolipids were noticeably increased in amounts during the course of neural differentiation of P19 EC cells induced by retinoic acid (RA, all- trans form). Applying neoglycolipid technology and in situ TLC-LSIMS, the oligosaccharide chains of these scarce Le x bearing glycolipids were partially characterized after released by endoglycoceramidase and subsequent conversion into neoglycolipids. In order to understand the enzymatic basis for the expression of Le x bearing glycolipids, we measured glycolipid, glycoprotein and oligosaccharide fucosyltransferase (Fuc-T) activities using appropriate substrates in P19 EC cells with or without RA treatment. All three Fuc-Ts were increased after RA treatment and the highest activity was in the differentiated neural cells. We then investigated the two possible Fuc-T genes that might be responsible for these changes using RT-PCR analysis. Mouse Fuc-TIX (mFuc-TIX) transcript was detected in all cell types but it was only strongly expressed in RA-induced aggregates and neural cells. In the case of mouse Fuc-TIV (mFuc-TIV) gene, its transcript was only detectable in RA-induced aggregates and not found in either undifferentiated or RA-induced neural cells. These results strongly support that RA induces only a transient expression of the mFuc-TIV gene in cell aggregates but a more persistent expression of the mFuc-TIX gene at the transcription level throughout neural cell differentiation. The mFuc-TIX gene is probably the main cause for the increased expression of Le x glycoconjugates during neural differentiation of P19 EC cells.