Glycosylation of CD4. Tunicamycin inhibits surface expression.
The T-cell surface glycoprotein CD4 plays an important role in mediating cellular immunity and serves as the receptor for human immunodeficiency virus. We have examined the glycosylation of CD4 and asked whether carbohydrate addition is essential for proper expression of the glycoprotein on the cell membrane. Under conditions where treatment of CD4+ human acute lymphoblastic leukemia cells (CEM-CM3 cells) with the glycosylation inhibitor tunicamycin decreased surface expression of CD4 in a time- and concentration-dependent manner, the surface expression of several other glycoproteins was unaffected. Incubation with tunicamycin for 48 h inhibited mannose incorporation by 98%, caused a 76% decrease in CD4 surface expression as judged by flow cytometry, and had little effect on methionine incorporation. Scatchard analysis showed a decrease in the total number of CD4 molecules on the cell surface from 17,000 to 8,900 after 24 h of tunicamycin treatment. Immunoprecipitation of metabolically labeled CD4 revealed the presence of an unglycosylated precursor in tunicamycin-treated cells. The observed difference between the Mr of the glycoprotein and its precursor is consistent with glycosylation at two potential N-linked sites. However, this precursor could not be detected by measuring steady state levels by immunoblotting. Also, no intracellular accumulation of CD4 in tunicamycin-treated cells was detectable using immunofluorescence microscopy. We conclude that surface expression of CD4 depends on glycosylation of the protein and that the unglycosylated precursor is preferentially degraded.
König
R
R
Laboratory of Biochemistry and Metabolism, National Institute of Diabetes, Digestive and Kidney Diseases, Bethesda, Maryland 20892.
Ashwell
G
G
Hanover
J A
JA
D000945:Antigens, Differentiation, T-Lymphocyte / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D007700:Kinetics; D007945:Leukemia, Lymphoid; D008358:Mannose / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000096:biosynthesis; D008715:Methionine / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000187:drug effects*; D014415:Tunicamycin / Q000494:pharmacology*
D000945:Antigens, Differentiation, T-Lymphocyte; D008562:Membrane Glycoproteins; D014415:Tunicamycin; D008715:Methionine; D008358:Mannose