Biosynthesis and intracellular transport of a membrane glycoprotein (plgp57) of the prelysosome compartment

Summary

We have characterized the biosynthesis and intracellular transport of a membrane glycoprotein, designated plgp57, which is found predominantly in the prelysosome compartment (PLC) of Madln-Darby bovine kidney cells. In pulse-chase experiments, plgp57 was found to be initially synthesized as a 35 kDa precursor which was modified to yield a diffuse ∼57 kDa mature form. Digestion with endoglycosidase H (endo H) demonstrated that the 35 kDa precursor contained three endo H-sensitive, high mannose-type oligosaccharides which became mofidifed to endo H-resistant, complex-type sugars on the ∼57 kDa mature form. Labelling cells in the presence of tunicamycin and treatment of the 35 kDa precursor with endo H revealed that plgp57 has a core protein of 24 kDa to which three N-asparagine-liked oligosaccharides are attached. Other experiments Indicated that plgp57 could be differentially glycosylated on a common 24 kDa core protein in different cell types. The half-lives of the glycosylated and non-glycosylated forms of plgp57 were ∼18 and ∼13 h, respectively. Glycosylated and non-glycosylated plgp57 exhibited similar steady-state intracellular distributions, indicating that targeting of plgp57 to the PLC does not require carbohydrate address markers. Pulse-labelling of cells followed by organelle fractionation at various chase times revealed a t_1/2 = ∼1 h for the transit of newly synthesized plgp57 to the PLC. Finally, amino terminal sequencing of plgp57 revealed the similarity of this protein to the CD63/ME491 family of membrane glycoproteins.

Key Words:

• Prelysosome compartment
• late endosome
• glycoprotein
• membrane trafficking
• CD63/ME491 family