Structure-function studies of the C3a-receptor: C-terminal serine and threonine residues which influence receptor internalization and signaling.

Abstract

The anaphylatoxic peptide C3a is a pro-inflammatory mediator generated during complement activation, whose specific G protein coupled receptor is expressed on granulocytes, monocytes, mast cells, activated lymphocytes, and in the nervous tissue. We have generated RBL-2H3 cell clones stably expressing mutants of the human C3a-receptor (C3aR) with combined alanine (Ala) substitutions of ten C-terminal serine (Ser) or threonine (Thr) residues, which may represent putative phosphorylation sites to characterize their role in ligand-induced C3aR internalization and signaling. Ser475/479 and Thr480/481 as well as Ser449 seemed not to be involved in ligand-induced receptor internalization. Either directly or by a conformational change they even "inhibit" C3aR internalization. In contrast, mutants with Ala substitutions at Ser465/470 and Thr463/466 were poorly internalized, and Thr463 seemed to be the most important C-terminal Thr or Ser residue directly effecting receptor internalization. However, it is likely that other C3aR regions additionally participate in this negative feed-back mechanism since even mutants with multiple Ala substitutions still internalized to a limited degree. Interestingly, in a mutant with a single exchange of Ser449 to Ala, the signal transduction assessed by a Ca(2+) assay and [(35)S]GTP gamma S-binding on HEK cells transiently co-transfected with G-alpha 16 or G-alpha O, respectively, was severely impaired, indicating that this residue of C3aR is involved in G protein coupling.

Cite this paper

@article{Settmacher2003StructurefunctionSO, title={Structure-function studies of the C3a-receptor: C-terminal serine and threonine residues which influence receptor internalization and signaling.}, author={Britta Settmacher and Claudia Rheinheimer and Henning Hamacher and Robert S. Ames and Alan Wise and Lesley Jenkinson and Daniel Bock and Myriam Schaefer and Joerg Koehl and Andreas Klos}, journal={European journal of immunology}, year={2003}, volume={33 4}, pages={920-7} }