Plasma membrane ectoglycosyltransferase activity of L1210 murine leukemic cells

✍ Scribed by R. J. Bernacki

• Publisher: John Wiley and Sons
• Year: 1974
• Tongue: English
• Weight: 780 KB
• Volume: 83
• Category: Article
• ISSN: 0021-9541
• DOI: 10.1002/jcp.1040830317

✦ Synopsis

Abstract

L1210 murine leukemia cells after treatment with Cl. perfringens neuraminidase at pH 7.0 incorporated six times more N‐acetylneuraminic acid‐[C^14^] than control cells when incubated for 30 minutes with cytidine 5′‐monophosphate N‐acetylneuraminic‐[C^14^] acid and three times more galactose‐[C^14^] when incubated with uridine diphosphate galactose‐[C^14^]. These sugars were incorporated in a 10% trichloracetic acid insoluble fraction and more than 75% of the incorporated N‐acetylneuraminic acid‐ [C^14^] could be removed by further treatment of these cells with neuraminidase. The incorporation of N‐acetylneuraminic acid‐ [C^14^] as a function of time was divided into two rates: a rapid one, active during the first 30 minutes followed by a slower one, similar to the rate observed with untreated cells. The addition of Ba^++^ and Ca^++^ ions at 8.3 mM increased the incorporation of N‐acetylneuraminic acid‐ [C^14^] by 25% while 8.3 mM EDTA decreased activity by 58% . The addition of Zn^++^ or Hg^++^ at similar concentrations abolished the incorporation almost completely. The optimal pH for the incorporation of N‐acetylneuraminic acid‐ [C^14^] by these neuraminidase treated cells was 6.5. These data suggest that ectoglycosyltransferases are present on the outer surface of the plasma membrane of L1210 cells and are able to catalyze the addition of radiolabeled nucleotide sugars onto macromolecular acceptors (cell surface glycoproteins and glycolipids) prepared by prior incubation of the cells with neuraminidase. Use of these procedures for labeling outer cell surfaces may also prove to be valuable for the study of plasma membrane glycoprotein and glycolipid structure, synthesis, and turnover.