Oligosaccharide reprocessing and recycling of a cell surface glycoprotein in cultured rat hepatocytes.
The metabolism of the cell surface glycoprotein dipeptidyl peptidase IV(DPPIV) was studied in cultured rat hepatocytes. In pulse-chase labelling experiments using L-[35S]methionine a 100-kDa high-mannose precursor polypeptide is converted into the mature complex-type 110-kDa glycoprotein. Digestion with exo- and endoglycosidases and metabolic labelling with radioactive sugars demonstrate that the 110-kDa form contains about 6 complex-type oligosaccharides which are fucosylated and sialylated. About 25 min after the beginning of the pulse-labelled glycoprotein appears in the sinusoidal membrane. Physiologically only the 110-kDa form is found in the cell surface. If cell surface DPP IV was desialylated by sialidase at 4 degrees C, it is resialylated during incubation at 37 degrees C. This oligosaccharide reprocessing indicates that the surface glycoprotein has been recycled to the cell compartment containing terminal glycosyltransferases (presumably the trans Golgi system). Two different methods demonstrate internalization of cell surface DPP IV: 1) The complex cell surface DPPIV -anti-DPP IV-antibody -L-[35S]methionine-labelled secondary goat-anti-mouse-antibody formed at 4 degrees C becomes less accessible to trypsin during incubation at 37 degrees C. 2) Part of the complex plasma membrane DPP IV-anti-DPP IV-antibody formed in the cold cannot be recognized by the radioactive secondary antibody after rewarming. Internalization is not blocked by inhibition of protein synthesis with cycloheximide. During internalization of plasma membrane DPP IV its concentration in the membrane remains constant.(ABSTRACT TRUNCATED AT 250 WORDS)
Kreisel
W
W
Medizinische Klinik, Klinikum der Albert-Ludwigs-Universität, Freiburg, Germany.
Hildebrandt
H
H
Mössner
W
W
Tauber
R
R
Reutter
W
W
D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D018819:Dipeptidyl Peptidase 4; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / Q000096:biosynthesis / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D006596:Hexosaminidases / Q000378:metabolism; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism*; D009439:Neuraminidase / Q000378:metabolism; D009844:Oligosaccharides / Q000378:metabolism*; D011233:Precipitin Tests; D051381:Rats
D008562:Membrane Glycoproteins; D009844:Oligosaccharides; D006596:Hexosaminidases; D009439:Neuraminidase; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases; D018819:Dipeptidyl Peptidase 4