Distinct functional sites for human immunodeficiency virus type 1 and stromal cell-derived factor 1alpha on CXCR4 transmembrane helical domains.
The entry of human immunodeficiency virus type 1 (HIV-1) into the cell is initiated by the interaction of the viral surface envelope protein with two cell surface components of the target cell, CD4 and a chemokine coreceptor, usually CXCR4 or CCR5. The natural ligand of CXCR4 is stromal cell-derived factor 1alpha (SDF-1alpha). Whereas the overlap between HIV-1 and SDF-1alpha functional sites on the extracellular domains of CXCR4 has been well documented, it has yet to be determined whether there are sites in the transmembrane (TM) helices of CXCR4 important for HIV-1 and/or SDF-1alpha functions, and if such sites do exist, whether they are overlapping or distinctive for the separate functions of CXCR4. For this study, by employing alanine-scanning mutagenesis, (125)I-SDF-1alpha competition binding, Ca(2+) mobilization, and cell-cell fusion assays, we found that the mutation of many CXCR4 TM residues, including Tyr(45), His(79), Asp(97), Pro(163), Trp(252), Tyr(255), Asp(262), Glu(288), His(294), and Asn(298), could selectively decrease HIV-1-mediated cell fusion but not the binding activity of SDF-1alpha. Phe(87) and Phe(292), which were involved in SDF-1alpha binding, did not play a significant role in the coreceptor activity of CXCR4, further demonstrating the disconnection between physiological and pathological activities of CXCR4 TM domains. Our data also show that four mutations of the second extracellular loop, D182A, D187A, F189A, and P191A, could reduce HIV-1 entry without impairing either ligand binding or signaling. Taken together, our first detailed characterization of the different functional roles of CXCR4 TM domains may suggest a mechanistic basis for the discovery of new selective anti-HIV agents.
Tian
Shaomin
S
Departments of Biochemistry, University of Illnois at Urbana-Champaign, 61801, USA.
Choi
Won-Tak
WT
Liu
Dongxiang
D
Pesavento
James
J
Wang
Youli
Y
An
Jing
J
Sodroski
Joseph G
JG
Huang
Ziwei
Z
D001665:Binding Sites; D002460:Cell Line; D054377:Chemokine CXCL12; D019743:Chemokines, CXC / Q000378:metabolism*; D015699:HIV Envelope Protein gp120 / Q000378:metabolism; D015497:HIV-1 / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D008958:Models, Molecular; D011485:Protein Binding; D017433:Protein Structure, Secondary; D019718:Receptors, CXCR4 / Q000378:metabolism / Q000502:physiology*; D017154:Stromal Cells; D014779:Virus Replication
C515255:CXCL12 protein, human; D054377:Chemokine CXCL12; D019743:Chemokines, CXC; D015699:HIV Envelope Protein gp120; D019718:Receptors, CXCR4