CHARACTERIZATION OF MUCINS AND PROTEOGLYCANS SYNTHESIZED BY A MUCIN-SECRETING HT-29 CELL SUBPOPULATION

Citation
G. Huet et al., CHARACTERIZATION OF MUCINS AND PROTEOGLYCANS SYNTHESIZED BY A MUCIN-SECRETING HT-29 CELL SUBPOPULATION, Journal of Cell Science, 108, 1995, pp. 1275-1285
Citations number
42
Categorie Soggetti
Cell Biology
Journal title
ISSN journal
00219533
Volume
108
Year of publication
1995
Part
3
Pages
1275 - 1285
Database
ISI
SICI code
0021-9533(1995)108:<1275:COMAPS>2.0.ZU;2-Q
Abstract
HT-29 cells selected by adaptation to 10(-5) M methotrexate (HT-29 MTX ) are a homogeneous cell population producing high amounts of mucin, I ntracellular mucins and proteoglycans were isolated from these cells b y ultracentrifugation of cell lysates on a cesium bromide gradient and further separated by anion-exchange high perfomance liquid chromatogr aphy. The major mucin fraction isolated was characterized by a high hy droxy amino acid content (40%), a Thr/Ser ratio of 1.52, a high sialic acid content, and a low sulfate content, When the same procedure was applied to undifferentiated HT-29 cells, a minor mucin fraction was is olated which appeared less sialylated and more sulfated, The major pro teoglycan species identified in HT-29 MTX cells showed less acidic beh avior than the proteoglycan isolated from HT-29 cells. The effect of b refeldin A and the sugar analog GalNAc-alpha-O-benzyl on the synthesis and biochemical properties of mucins synthesized by HT-29 MTX cells w as examined, Brefeldin A induced the synthesis of more-sulfated mucins , GalNAc-alpha-O-benzyl treatment resulted in mucins with an increased content of T antigen and a 13-fold lower sialic acid content, We show that GalNAc-alpha-O-benzyl was metabolized by the cells to Gal beta 1 -3GalNAc-alpha-O-benzyl, which, in turn, was a potent competitive inhi bitor of the O-glycan alpha-2,3-sialyltransferase. These results illus trate the suitability of HT-29 MTX cells as a model to analyse mucin s ynthesis and sialylation.