PT  - JOURNAL ARTICLE
AU  - Satya P Singh
AU  - John F Foley
AU  - Hongwei H Zhang
AU  - Darrell E Hurt
AU  - Jennifer L Richards
AU  - Craig S Smith
AU  - Fang Liao
AU  - Joshua M Farber
TI  - Selectivity In The Use of Gi/o Proteins Is Determined by the DRF Motif In CXCR6 And Is Cell-type Specific
AID  - 10.1124/mol.115.099960
DP  - 2015 Jan 01
TA  - Molecular Pharmacology
PG  - mol.115.099960
4099  - http://molpharm.aspetjournals.org/content/early/2015/08/27/mol.115.099960.short
4100  - http://molpharm.aspetjournals.org/content/early/2015/08/27/mol.115.099960.full
AB  - CXCR6, the receptor for CXCL16, is expressed on multiple cell types and can be a co-receptor for HIV-1. Except for CXCR6, all human chemokine receptors contain the D3.49R3.50Y3.51 sequence, and all but two contain A3.53, at the cytoplasmic terminus of the third transmembrane helix (H3C), a region within class A G-protein-coupled-receptors that contacts G proteins. In CXCR6, H3C contains D3.49R3.50F3.51I3.52V3.53 at positions 126-130. We investigated the importance and inter-dependence of the canonical D126 and the non-canonical F128 and V130 in CXCR6 by mutating D126 to Y, F128 to Y, and V130 to A, singly and in combination. For comparison, we mutated the analogous positions, D142, Y144, and A146 to Y, F, and V, respectively, in CCR6, a related receptor containing the canonical sequences. Mutants were analyzed in both HEK-293T and Jurkat E6-1 cells. Our data show that for CXCR6 and/or CCR6, mutations in H3C can affect both receptor signaling and chemokine binding; non-canonical H3C sequences are functionally linked, with dual changes mitigating effects of single mutations; mutations in H3C that compromise receptor activity show selective defects in the use of individual Gi/o proteins; and the effects of mutations in H3C on receptor function and selectivity in Gi/o protein use can be cell-type specific. Our findings indicate that the ability of CXCR6 to make promiscuous use of the available Gi/o proteins is exquisitely dependent on sequences within the H3C, and suggest that the native sequence allows for preservation of this function across different cellular environments.