title: C5aR expression in a novel GFP reporter gene knockin mouse: implications for the mechanism of action of C5aR signaling in T cell immunity.
abstract: C5aR is a G protein-coupled receptor for the anaphylatoxin C5a and mediates many proinflammatory reactions. C5aR signaling also has been shown to regulate T cell immunity, but its sites and mechanism of action in this process remain uncertain. In this study, we created a GFP knockin mouse and used GFP as a surrogate marker to examine C5aR expression. GFP was knocked into the 3'-untranslated region of C5ar1 by gene targeting. We show that GFP is expressed highly on Gr-1(+)CD11b(+) cells in the blood, spleen, and bone marrow and moderately on CD11b(+)F4/80(+) circulating leukocytes and elicited peritoneal macrophages. No GFP is detected on resting or activated T lymphocytes or on splenic myeloid or plasmacytoid dendritic cells. In contrast, 5-25% cultured bone marrow-derived dendritic cells expressed GFP. Interestingly, GFP knockin prevented cell surface but not intracellular C5aR expression. We conclude that C5aR is unlikely to play an intrinsic role on murine T cells and primary dendritic cells. Instead, its effect on T cell immunity in vivo may involve CD11b(+)F4/80(+) or other C5aR-expressing leukocytes. Further, our data reveal a surprising role for the 3'-untranslated region of C5aR mRNA in regulating C5aR protein targeting to the plasma membrane.
authors.affiliation: Department of Pharmacology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
authors.forename: Jason
authors.initials: J
authors.lastname: Dunkelberger
authors.forename: Lin
authors.initials: L
authors.lastname: Zhou
authors.forename: Takashi
authors.initials: T
authors.lastname: Miwa
authors.forename: Wen-Chao
authors.initials: WC
authors.lastname: Song
mesh_terms: D020413:3' Untranslated Regions / Q000235:genetics; D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology / Q000276:immunology*; D039481:CD11b Antigen / Q000276:immunology; D002465:Cell Movement / Q000276:immunology; D005260:Female; D015870:Gene Expression; D055879:Gene Knock-In Techniques; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000276:immunology; D007111:Immunity, Cellular / Q000235:genetics / Q000276:immunology*; D008264:Macrophages / Q000166:cytology / Q000276:immunology*; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D044087:Receptor, Anaphylatoxin C5a / Q000235:genetics* / Q000276:immunology; D015398:Signal Transduction; D013154:Spleen / Q000166:cytology / Q000276:immunology*; D013601:T-Lymphocytes / Q000166:cytology / Q000276:immunology*
chemical_list: D020413:3' Untranslated Regions; D039481:CD11b Antigen; D044087:Receptor, Anaphylatoxin C5a; D049452:Green Fluorescent Proteins