PMID- 10700388 OWN - NLM STAT- MEDLINE DCOM- 20000412 LR - 20071114 IS - 0003-9861 (Print) IS - 0003-9861 (Linking) VI - 375 IP - 2 DP - 2000 Mar 15 TI - alpha(1,3)fucosyltransferases expressed by the gain-of-function Chinese hamster ovary glycosylation mutants LEC12, LEC29, and LEC30. PG - 322-32 AB - Gain-of-function glycosylation mutants provide access to glycosylation pathways, glycosylation genes, and mechanisms that regulate expression of a glycotype. Previous studies have shown that the gain-of-function Chinese hamster ovary (CHO) mutants LEC12, LEC29, and LEC30 express an N-ethylmaleimide-resistant alpha(1, 3)fucosyltransferase (alpha(1,3)Fuc-T) activity that is not detected in CHO cells and that generates the Lewis(X) but not the sialyl-Lewis(X) determinant. The three mutants differ, however, in lectin resistance properties, expression of fucosylated antigens, and in vitro alpha(1,3)Fuc-T activities. In this paper we show that each mutant expresses Fuc-TIX, but only LEC30 cells express Fuc-TIV. Using genomic PCR and reverse-transcriptase (RT)-PCR strategies, we isolated coding portions of the CHO Fut4 and Fut9 genes. Each gene is present in a single copy in the CHO and mutant genomes. The Fut4 gene is expressed only in LEC30 cells, while all three mutants express the Fut9 gene. Interestingly, the fucosylation phenotypes of LEC12 and LEC29 cells do not correlate with the relative abundance of their Fut9 gene transcripts (LEC29 >> LEC12). Compared to LEC29 cells, LEC12 cells have an approximately 40-fold higher in vitro alpha(1,3)Fuc-T activity and bind the VIM-2 monoclonal antibody, whereas LEC29 cells do not bind VIM-2. Mixing experiments did not detect Fuc-TIX inhibitory activity in LEC29 cell extracts, and CHO cells expressing a transfected Fut9 gene behaved like LEC12 cells. Therefore, it seems that LEC29 cells may not translate their more abundant Fut9 gene transcripts efficiently or may not synthesize appropriate acceptors for internal alpha(1,3)fucosylation. Alternatively, LEC12 cells may possess, in addition to Fuc-TIX, a novel alpha(1,3)Fuc-T activity. CI - Copyright 2000 Academic Press. FAU - Patnaik, S K AU - Patnaik SK AD - Department of Cell Biology, Albert Einstein College of Medicine, New York, New York, 10461, USA. FAU - Zhang, A AU - Zhang A FAU - Shi, S AU - Shi S FAU - Stanley, P AU - Stanley P LA - eng SI - GENBANK/AF230460 GR - P01 13330/PHS HHS/United States GR - R01 36434/PHS HHS/United States PT - Journal Article PT - Research Support, Non-U.S. Gov't PT - Research Support, U.S. Gov't, P.H.S. PL - United States TA - Arch Biochem Biophys JT - Archives of biochemistry and biophysics JID - 0372430 RN - 0 (Antibodies, Monoclonal) RN - 0 (DNA, Complementary) RN - 0 (Epitopes) RN - 0 (RNA, Messenger) RN - EC 2.4.1.- (Fucosyltransferases) RN - EC 2.4.1.152 (galactoside 3-fucosyltransferase) SB - IM MH - Amino Acid Sequence MH - Animals MH - Antibodies, Monoclonal/immunology MH - Blotting, Northern MH - CHO Cells MH - Cloning, Molecular MH - Cricetinae MH - DNA, Complementary/genetics MH - Epitopes/immunology MH - Fucosyltransferases/chemistry/*genetics/immunology/*metabolism MH - *Gene Expression MH - Glycosylation MH - Molecular Sequence Data MH - Molecular Weight MH - Mutation/*genetics MH - Open Reading Frames/genetics MH - Phenotype MH - Phylogeny MH - RNA, Messenger/analysis/genetics MH - Transfection EDAT- 2000/03/04 09:00 MHDA- 2000/04/15 09:00 CRDT- 2000/03/04 09:00 PHST- 2000/03/04 09:00 [pubmed] PHST- 2000/04/15 09:00 [medline] PHST- 2000/03/04 09:00 [entrez] AID - S0003-9861(99)91693-4 [pii] AID - 10.1006/abbi.1999.1693 [doi] PST - ppublish SO - Arch Biochem Biophys. 2000 Mar 15;375(2):322-32. doi: 10.1006/abbi.1999.1693.