Identification of the TRIM5 specificity domain through evolutionary analysis:

My studies of the primate TRIM5a protein illustrate the power of this approach. TRIM5a interacts with and disrupts the capsids of retroviruses uncoating in the cytoplasm of host cells. Depending on the ortholog encoded by a specific host, this protein has activity against diverse retroviruses including HIV, but is unfortunately inactive against HIV in humans. I sequenced this gene from a panel of primates and, using statistical methods, identified a small patch of 13 amino acid sites (out of 400) that has been the direct target of positive selection. I predicted and then proved that this patch represents the anti-viral interface and accounts for the specificity of TRIM5a restriction. Through amino acid substitution in this domain, I was able to convey HIV recognition to the human TRIM5a protein in vitro. Thus, I was able to bypass standard biochemical dissection of this protein by examining evolutionary signatures and then performing a single theory-based experiment to identify the virus-binding motif.

Identification of human polymorphism relevant to viral infection:

One enigma that surrounds the evolution of immunity genes is how they are selectively maintained through eras of low pathogenic challenge. In fact, modern humans currently have a very low retroviral load. We are known to be susceptible to only two families of exogenous retroviruses (HIV and HTLV) and have largely or completely inactivated our substantial endogenous retroviral load. I was interested in knowing how TRIM5 has fared through this era of low retroviral challenge, and if this history contributes to our current susceptibility to HIV. To answer this question, I sequenced and characterized TRIM5 variation in a world-wide sample of human genomes. I found six coding SNPs and made TRIM5 constructs containing each of these amino acid changes. In collaboration with a retrovirology lab, I demonstrated that one of these SNPs (H43Y) is broadly detrimental to TRIM5 anti-viral function. Furthermore, this allele is rare in the Old World (4%), but common in the Americas (43%). I proposed that this impaired allele drifted to high frequency after human migration to the New World because the loss of function did not provide a contemporary fitness disadvantage. This indicates that TRIM5 has not been strictly maintained in the last 10,000 years. I proposed that there may be future health implications of this impaired retroviral restriction allele segregating at high frequency in the human population.

Identification of novel retroviral restriction genes in primates:

I demonstrated that two primate genes encoding known retroviral defense proteins with anti-HIV activity (TRIM5 and APOBEC3G) show some of the most intense signals of positive selection so far observed. My next goal was to turn the tables and use signatures of positive selection to identify new anti-viral genes. First, I identified other genes in the APOBEC gene family that are evolving under positive selection and predicted that these encode additional restriction factors, predictions which were subsequently verified by other groups. I then analyzed the human TRIM gene family for positive selection. Surprisingly, I find that positive selection is more rare in this family, with the startling exception of a new expansion of TRIM genes that has occurred so recently in humans that the gene cluster is still polymorphic in our population. Positive selection of these genes, together with their young age and similarity to TRIM5, evokes the hypothesis that they are currently engaged in an evolutionary "arms race" with an unknown retroviral pathogen.