Role of CXCR3 carboxyl terminus and third intracellular loop in receptor-mediated migration, adhesion and internalization in response to CXCL11.
The chemokine receptor CXCR3 is predominantly expressed on activated T and natural killer (NK) cells. CXCR3 and its ligands, CXCL11, CXCL10, and CXCL9, play a major role in T-helper 1 (Th1)-dependent inflammatory responses. CXCL11 is the most dominant physiological inducer of adhesion, migration, and internalization of CXCR3. To study the role of CXCR3 carboxyl-terminus and the third intracellular (3i) loop in chemokine-mediated migration, adhesion, and CXCR3 internalization, we generated CXCR3 receptors mutated in their distal (Ser-Thr domain) or proximal (trileucine domain) membrane carboxyl terminus, and/or the third intracellular loop. We found that migration of CXCR3-expressing HEK 293 cells toward CXCL11 was pertussis toxin-dependent and required the membrane proximal carboxyl terminus of CXCR3. Internalization induced by CXCL11 and protein kinase C (PKC) activation was also regulated by the membrane proximal carboxyl terminus; however, only CXCL11-induced internalization required the LLL motif of this region. Internalization and Ca(2+) flux induced by CXCL11 were independent of the 3i loop S245, whereas migration at high CXCL11 concentrations, integrin-dependent adhesion, and actin polymerization were S245 dependent. Our findings indicate that CXCL11-dependent CXCR3 internalization and cell migration are regulated by the CXCR3 membrane proximal carboxyl terminus, whereas adhesion is regulated by the 3i loop S245. Thus, distinct conformational changes induced by a given CXCR3 ligand trigger different downstream effectors of adhesion, motility, and CXCR3 desensitization.
Dagan-Berger
Michal
M
Gene Therapy Institute, Hadassah University Hospital, PO Box 12000, Jerusalem, Israel.
Feniger-Barish
Rotem
R
Avniel
Shani
S
Wald
Hanna
H
Galun
Eithan
E
Grabovsky
Valentin
V
Alon
Ronen
R
Nagler
Arnon
A
Ben-Baruch
Adit
A
Peled
Amnon
A
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling; D002448:Cell Adhesion / Q000502:physiology*; D002460:Cell Line; D002465:Cell Movement / Q000502:physiology*; D054371:Chemokine CXCL11; D019743:Chemokines, CXC / Q000502:physiology*; D017931:DNA Primers; D005353:Fibronectins / Q000502:physiology; D018014:Gene Transfer Techniques; D006801:Humans; D007668:Kidney; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010957:Plasmids; D054367:Receptors, CXCR3; D019707:Receptors, Chemokine / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D011994:Recombinant Proteins / Q000378:metabolism; D014162:Transfection; D019010:Vascular Cell Adhesion Molecule-1 / Q000502:physiology
C515202:CXCL11 protein, human; C515187:CXCR3 protein, human; D054371:Chemokine CXCL11; D019743:Chemokines, CXC; D017931:DNA Primers; D005353:Fibronectins; D054367:Receptors, CXCR3; D019707:Receptors, Chemokine; D011994:Recombinant Proteins; D019010:Vascular Cell Adhesion Molecule-1; D002118:Calcium