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Effect of p58GTA on β-1,4-galactosyltransferase 1 activity and cell-cycle in human hepatocarcinoma cells

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Abstract

β-1,4-galactosyltransferase 1 (β1,4-GT 1) is the key enzyme transferring galactose to the terminal N-acetylglucosamine (GlcNAc) forming Galβ1→4GlcNAc structure in the Golgi apparatus. In addition, it also serves as a cell adhesion molecule by recognizing and binding to terminal GlcNAc of glycoconjugates on the adjacent cell surface and matrix through a subpopulation of the enzyme distributed on the cell surface. Transient expression of the p58GTA protein kinase, which belongs to the p34cdc2-related supergene family, could enhance β1,4-GT 1 total activity in COS cells. In this study, the p58GTA interaction with β1,4-GT 1 was confirmed using an in vitro assay with the TNT® Coupled Reticulocyte Lysate System. An expression vector containing p58GTA was stably transfected into 7721 cells, a human hepatocarcinoma cell line, expression was confirmed by Northern and Western blot analyses. The cells transfected with p58GTA (p58GTA/7721) contained 1.9 times higher total β1,4-GT 1 activity and 2.6 times higher cell-surface β1,4-GT 1 activity than the mock transfected cells (pcDNA3/7721). However, Ricinus communis agglutinin-I lectin blot analysis revealed that the enhanced β1,4-GT 1 activity did not increase the Galβ1→4GlcNAc groups on most of the membrane proteins in p58GTA/7721 cells. By flow cytometry analysis, it was found that the p58GTA/7721 cells were G2/M phase arrested, compared with the pcDNA3/7721 cells. These results suggest that the p58GTA stable transfection into human hepatocarcinoma cells could enhance the two β1,4-GT 1 subcellular pool activities independently and change its cell-cycle without modifying the β-1,4-linked galactose residues on most membrane proteins.

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Authors and Affiliations

  1. Gene Research Center, Medical Center of Fudan University (Former Shanghai Medical University), Shanghai, P.R. China, 200032

    Song W. Zhang, Song L. Xu, Ming M. Cai, Jun Yan, Xiao Y. Zhu, Yun Hu & Jian X. Gu

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  1. Song W. Zhang
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  2. Song L. Xu
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  3. Ming M. Cai
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  7. Jian X. Gu
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Correspondence to Jian X. Gu.

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Zhang, S.W., Xu, S.L., Cai, M.M. et al. Effect of p58GTA on β-1,4-galactosyltransferase 1 activity and cell-cycle in human hepatocarcinoma cells. Mol Cell Biochem 221, 161–168 (2001). https://doi.org/10.1023/A:1010932211745

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