Esposito, M. to the control Caucasian men (= 0.02). This increase was mostly attributable to a higher frequency of the ?2459 A/G versus the ?2459 A/A genotype in individuals heterozygous for the 32 allele (= 0.012). No protective influence of the CCR2-64I allele was observed. The haplotypes CCR5 ORF 32/CCR5 E3 ligase Ligand 10 ?2459A (in complete linkage disequilibrium) and CCR5 ORF wt/CCR5 ?2459G had a cumulative negative effect on the expression of CCR5, since we measured significantly reduced CCR5 densities on both T-helper cells and monocytes only when both haplotypes were present. Densities of CCR5 on lymphocytes and monocytes were correlated (= 0.59; 0.0001), indicating concordance of CCR5 expression patterns across different cell types. We conclude that the CCR5 ORF Rabbit polyclonal to HYAL2 32/wt-CCR5 ?2459 A/G genotype combination offers an advantage in resisting sexual HIV-1 transmission and that this effect is mediated by a relative paucity of CCR5 on potential target cells of HIV-1. Human immunodeficiency virus type 1 (HIV-1) requires expression of CD4 in conjunction with a chemokine coreceptor, most frequently CCR5 or CXCR4, to productively infect target cells. Since the discovery of these coreceptors, it has become well established that alterations in their gene expression and function can impact HIV-1 disease progression. For example, inheritance of the chemokine receptor (CR) polymorphisms CCR5 open reading frame (ORF) 32, CCR5 promoter ?2459 A-to-G, and CCR2 ORF 190 G-to-A (CCR2-64I) are associated with delayed development of the AIDS in HIV-1-infected patients (4, 5, 9, 14, 16, 17, 20, 21, 29, 33, 35, 36, 38, 39, 45, 53, 54). Moreover, individuals homozygous for the CCR5-32 allele, comprising about 1% of the Caucasian population, have strongly reduced susceptibility to R5-dependent HIV-1 infection (7, 26, 49). However, it is clear that the infrequent inheritance of homozygosity for the CCR5-32 allele cannot account for the majority of persons worldwide who are repeatedly exposed to HIV-1 by high-risk activities but remain seronegative and uninfected. In this study, we investigated the contributions of the CCR5 ?2459G and CCR2-64I polymorphisms, as well as heterozygosity for CCR5-32, to protection against HIV-1 infection in a well-defined exposed seronegative (ES) longitudinal cohort (8, 12) in contrast to a low-risk uninfected control group. Whereas CCR5 ORF 32/32 homozygosity occurs in just 1% of Caucasians, the CCR5 ?2459 A/G plus CCR5 ORF 32/wt or CCR2 ORF 64I/wt genotype combinations comprise approximately 5% each. However, the effects of these two genotype combinations on the susceptibility to HIV-1 infection in vivo have not been conclusively determined (6, 7, 13, 14, 17, 22, 27, 30, 31, 37, 49, 53, 55, 61). Here, we have compared the frequencies of the CCR5 ORF wt/32, CCR5 ?2459A/G, and CCR2 ORF 64V/I alleles and allele combinations between the ES and the low-risk control group. In addition, we determined the impact of inheriting specific haplotypic combinations of these alleles in our cohorts on the expression of CCR5 on HIV-1 target cells. Our results indicate that inheritance of the CCR5 ORF 32/wt plus ?2459 A/G genotype combination occurs more commonly in ES E3 ligase Ligand 10 than in low-risk control individuals and that these genotypes together may act to reduce target cell susceptibility to HIV-1. MATERIALS AND METHODS Study populations. Ninety-three healthy HIV-1-seronegative individuals 18 years old who reported high-risk sexual activity with known HIV-1-infected partners were recruited within metropolitan Seattle for longitudinal study at the Fred Hutchinson Cancer Research Center HIV-1 Vaccine Trials Unit. Enrollment criteria were unprotected sexual intercourse with a known HIV-1-infected person 6 times in the previous 6 months or an average of twice weekly 4 months within 2 years of enrollment. Prior receipt of an HIV-1 vaccine was an exclusion criterion. Volunteers entering the study were designated ES. A medical history, physical examination, complete blood cell count, T-cell subset analysis, and HIV-1 testing were done at the screening visit to confirm eligibility. HIV-1 infection was excluded by E3 ligase Ligand 10 the following tests done at the screening visit and study day 0: E3 ligase Ligand 10 HIV-1/2 enzyme-linked immunosorbent assay (ELISA) and Western blot (50), HIV-1 plasma RNA reverse transcription-PCR (Amplicor HIV-1 Monitor; Roche Molecular Systems) E3 ligase Ligand 10 (19), and peripheral blood mononuclear cell (PBMC) DNA PCR (41). HIV-1/2 ELISA and Western blot were repeated every 3 months, and HIV-1 PBMC.