Endocytosis by the asialoglycoprotein receptor is independent of cytoplasmic serine residues.
The human asialoglycoprotein (ASGP) receptor, like most other plasma membrane receptors, has previously been shown to be phosphorylated at serine residues within the cytoplasmic domain. Phorbol esters, which activate protein kinase C, cause hyperphosphorylation and down-regulation of the ASGP receptor in HepG2 cells. To test the importance of serine residues for receptor traffic and function, we have mutated all the cytoplasmic serines of the two receptor subunits H1 (at positions 16 and 37) and H2 (at positions 12, 13, and 55) to alanines or glycines. Stable transfected fibroblast cell lines expressing either mutant H1 alone or both mutant subunits together were created and compared to cell lines expressing the respective wild-type proteins. Mutant and wild-type subunits were found to have very similar distributions between the cell surface and intracellular compartments. Constitutive internalization of H1 alone and ligand uptake and degradation by cells expressing both receptor subunits were not affected by the mutations. Cytoplasmic serines and serine phosphorylation are thus not essential for receptor function and intracellular traffic. Analysis of individual serine mutations identified serine-12 of subunit H2 as the major site of phosphorylation in the ASGP receptor.
Geffen
I
I
Department of Biochemistry, University of Basel, Switzerland.
Fuhrer
C
C
Spiess
M
M
D037263:Asialoglycoprotein Receptor; D001212:Asialoglycoproteins / Q000378:metabolism*; D001483:Base Sequence; D003593:Cytoplasm / Q000648:ultrastructure; D004252:DNA Mutational Analysis; D004705:Endocytosis; D006801:Humans; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000737:chemistry; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D010768:Phosphoserine / Q000737:chemistry; D011971:Receptors, Immunologic / Q000378:metabolism*; D013329:Structure-Activity Relationship; D014162:Transfection
D037263:Asialoglycoprotein Receptor; D001212:Asialoglycoproteins; D009841:Oligonucleotides; D010750:Phosphoproteins; D011971:Receptors, Immunologic; D010768:Phosphoserine