This project's focus is on two infectious diseases that continue to have tremendous impact on global health. Human Immunodeficiency Virus (HIV) is pandemic and Hepatitis B Virus (HBV) infection and HBV-associated hepatocellular carcinoma (HCC) is prevalent in East Asia, globally affecting millions of people and having no cure. The objective of this project is to identify host factors that contribute to the occurrence and development of these, and potentially other, infectious diseases. We aim to identify host genetic factors that affect host innate restriction or susceptibility in acquisition, replication, and pathogenesis of viral pathogens, and carcinogenesis, the mechanisms of which are not fully understood. The identification of host proteins involved in viral replication, in innate or acquired immunity, or in carcinogenesis pathways will provide critical insights for the rational development of antiviral drugs and effective vaccines. Our strategy is to search for genetic variants that differentially affect rates of infection, or the course of pathogenesis, and which thereby identify the variant-containing gene as conferring restriction or susceptibility to infection or progression. We are using both targeted gene and genome wide association study (GWAS) approaches, including Illumina and Affymetrix chip technologies, to discover genes associated with HIV-1, HBV infection and HBV-associated liver cancer. Accomplishments 1) HIV-1 resistant APOBEC3 (A3) genes. Six of the seven A3 genes (including A3B, A3G and A3F) are human innate resistance factors that confer resistance to HIV-1 by causing hypermutation of viral RNA. Previously we discovered genetic factors in the APOBEC3G and CUL5 pathway that affect AIDS progression. The seven APOBEC3 genes (A-H) are clustered in a 100 kb region in chromosome 22. We assessed the influence of the genetic variants and haplotype in all APOBEC3 genes (A-H) on HIV-1 infection and disease courses. Besides the associations we previous found in A3G and A3B, we have identified A3F genetic variants that retard AIDS progression. In vitro, APOBEC3F strongly inhibits HIV-1 and is partially resistant to HIV-1 vif, unlike APOBEC3G. We found that codon-changing variants rs5750728 (intronic or codon-changing 78 V/A in different isoforms) and 231 I/V were associated with protection from progression to AIDS in European Americans. We found that rs5750728 alters binding to nuclear proteins in HeLa cells, suggesting a regulator role. In collaboration with researchers at Johns Hopkins University, the interaction of the APOBEC3F with HIV-1 vif was evaluated; the carrying codon-changing SNP 231 I/V was shown to confer stronger resistance to vif that degrades A3F, possibly leading to more abundant A3F to inhibit HIV-1. It is of interest that in the alignment of A3F and vif sequences, the A3F region carrying 231 I/V appears to either be mimicked by or has co-evolved with HIV-1 vif. This may reveal a mechanism allowing HIV-1 to evade A3F by adapting to host A3F sequences. 2) ZNRD1 and HIV-1 infection. We investigated the effect of the SNPs in the ZNRD1 region on HIV-1 infection and progression in five U.S-based HIV-1 longitudinal cohorts. A haplotype in the ZNRD1 gene showed significant association with host restriction to HIV-1 acquisition. The functional relevance of the associated variant was demonstrated in a gene reporter assay that showed one promoter variant (rs3132130) increased the ZNRD1 gene expression. Further, in a DNA-protein interaction assay, the variant also confers differential allele-specific binding to nuclear proteins. Our findings provide novel evidence of significant roles of ZRND1 in modulating HIV/AIDS. This work has been published in Journal of Infectious Diseases, June 2014. 3) GWAS of HBV and HCC. HBV infection is extremely prevalent in China (HBsAg carrier rate was 7% in 2006). Many HBV infected patients develop cirrhosis and hepatocellular carcinoma (HCC). MGE has an ongoing collaboration with Dr. Zeng Zheng, Beijing University First Hospital, for genetic studies in a Chinese Hepatitis B virus infection cohort. This HBV cohort, initiated by MGE, is collaboration between 13 major institutes in China and MGE. The HBV cohort comprises the full spectrum of the HBV outcomes: HBV resistance, HBV clearance, chronic hepatitis, cirrhosis and HCC, making this the most comprehensive sample set covering natural HBV outcomes available. Our GWAS was performed on 1200 samples using the affymetrix SNP6.0 chip. We have completed quality control of genotyping calls and performed GWAS association analyses. The genotypes from 6 top associated SNPs were verified using a second TaqMan assay, and the association signals were validated in expanded sample sets in the same cohort. The GWAS identified HLA-DPA1 and HLA-DPB1 region as major genetic factors for HBV clearance and infection that passed the genome wide significance threshold. These results are in agreement of GWAS results from other groups as well as our previous published finding, in which HLA-DPA1 rs3077 was associated with not only reduced risk to HBV infection but also HBV clearance. We detected an association with host resistance to HBV infection in the HLA-DQ1 region that passed the genome wide significance (p5e-8). The top HLA-DQ1 SNP is an eQTL associated with HLA-DQ expression level in two published genome wide QTL studies. 5) We contributed genotype-phenotype data from the HIV/AIDS GWAS to the International for the Genomics of HIV to identify common variants associated with HIV acquisition. The conclusion of the study, published in PLoS Pathogens, was that genetic influences on HIV acquisition are either rare or have small effects. 6) We have initiated a longitudinal study of differential methylation in PBMCs from HIV-infected subjects at different stages of disease, comparing pre-infection samples from each subject with samples from the same subject at later stages of disease. We ran 82 samples from the DCG cohort on the Illumina Infinium 450 methylation chip. We are collaborating with the CCR bioinformatics group (CCR-IFX) to analyze the chip data. Quality of the reads was confirmed by the ability of the methylation data to predict the CD4 count of individuals, as confirmed by measured CD4 counts. Initial results show approximately 150 sites differentially methylated between pre- and post-infection samples, after correcting for age, CD4 count, and other confounding variables. 7) The Longitudinal Studies of Ocular Complications of AIDS (LSOCA) comprises 2393 patients with AIDS and with detailed follow-up clinical data to monitor incidence of ocular complications of AIDS and to determine the effect of antiretroviral treatment on immune function on the risk of ocular disease. We have genotyped this cohort for 15 candidate genes associated with age-related macular degeneration and for APOL1 variants for association with retinal arteriole injury and renal disease; analysis is ongoing.