Expression of glycoconjugates bearing the Lewis X epitope during neural differentiation of P19 EC cells.
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.
Osanai
T
T
Department of Biochemical Cell Research, Tokyo Metropolitan Institute of Medical Science, Tokyo, Japan. osanai@rinshoken.or.jp
Chai
W
W
Tajima
Y
Y
Shimoda
Y
Y
Sanai
Y
Y
Yuen
C T
CT
D000818:Animals; D015153:Blotting, Western; D002240:Carbohydrate Sequence; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D004790:Enzyme Induction / Q000187:drug effects; D000939:Epitopes / Q000096:biosynthesis* / Q000737:chemistry / Q000276:immunology; D005647:Fucosyltransferases / Q000235:genetics / Q000378:metabolism; D006017:Glycolipids / Q000032:analysis / Q000737:chemistry; D016256:Lewis X Antigen / Q000096:biosynthesis* / Q000737:chemistry / Q000276:immunology; D013058:Mass Spectrometry; D051379:Mice; D008969:Molecular Sequence Data; D009474:Neurons / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D009844:Oligosaccharides / Q000032:analysis / Q000737:chemistry; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014212:Tretinoin / Q000494:pharmacology
D000939:Epitopes; D006017:Glycolipids; D016256:Lewis X Antigen; D009844:Oligosaccharides; D012333:RNA, Messenger; D014212:Tretinoin; D005647:Fucosyltransferases; C031692:galactoside 3-fucosyltransferase