A comprehensive mapping of interactions among Epstein-Barr virus (EBV) proteins and interactions of EBV proteins with human proteins should provide specific hypotheses and a broad perspective on EBV strategies for replication and persistence. Interactions of EBV proteins with each other and with human proteins were assessed by using a stringent high-throughput yeast two-hybrid system. Overall, 43 interactions between EBV proteins and 173 interactions between EBV and human proteins were identified. EBV-EBV and EBV-human protein interaction, or ''interactome'' maps provided a framework for hypotheses of protein function. For example, LF2, an EBV protein of unknown function interacted with the EBV immediate early R transactivator (Rta) and was found to inhibit Rta transactivation. From a broader perspective, EBV genes can be divided into two evolutionary classes, ''core'' genes, which are conserved across all herpesviruses and subfamily specific, or ''noncore'' genes. Our EBV-EBV interactome map is enriched for interactions among proteins in the same evolutionary class. Furthermore, human proteins targeted by EBV proteins were enriched for highly connected or ''hub'' proteins and for proteins with relatively short paths to all other proteins in the human interactome network. Targeting of hubs might be an efficient mechanism for EBV reorganization of cellular processes.herpesvirus ͉ interactome ͉ replication ͉ yeast two hybrid

Human TRIM5␣ (TRIM5␣ hu ) only modestly inhibits human immunodeficiency virus type 1 (HIV-1) and does not inhibit simian immunodeficiency virus (SIV mac ). Alteration of arginine 332 in the TRIM5␣ hu B30.2 domain to proline, the residue found in rhesus monkey TRIM5␣, has been shown to create a potent restricting factor for both HIV-1 and SIV mac. Here we demonstrate that the potentiation of HIV-1 inhibition results from the removal of a positively charged residue at position 332 of TRIM5␣ hu. The increase in restricting activity correlated with an increase in the ability of TRIM5␣ hu mutants lacking arginine 332 to bind HIV-1 capsid complexes. A change in the cyclophilin A-binding loop of the HIV-1 capsid decreased TRIM5␣ hu R332P binding and allowed escape from restriction. The ability of TRIM5␣ hu to restrict SIV mac could be disrupted by the presence of any charged residue at position 332. Thus, charged residues in the v1 region of the TRIM5␣ hu B30.2 domain can modulate capsid binding and restriction potency. Therapeutic strategies designed to neutralize arginine 332 of TRIM5␣ hu might potentiate the innate resistance of human cells to HIV-1 infection.Primates express dominant restriction factors that block retrovirus infection soon after entry but prior to reverse transcription (1, 2, 5). Genetic studies of virus variants and restriction factor competition studies indicate that the viral capsid is the determinant of susceptibility to restriction (3, 3b, 6, 13, 14). Most early restriction in primates is mediated by TRIM5␣ (7,10,16,23,26). TRIM5␣ is a member of the tripartite motif family of proteins and contains RING, B-box 2, and coiled-coil (RBCC) domains (17). TRIM5␣ also contains a C-terminal B30.2/SPRY domain, which is required for retroviral restriction (23). Deletion of the B30.2 domain disrupts the ability of the TRIM5␣ protein to bind viral capsid complexes (20,24). Differences in the B30.2 domains of TRIM5␣ proteins from distinct primate species account for patterns of retrovirus restriction. For example, the rhesus monkey TRIM5␣ (TRIM5␣ rh ) potently blocks human immunodeficiency virus type 1 (HIV-1), which is only weakly inhibited by human TRIM5␣ (TRIM5␣ hu ) (23). Neither TRIM5␣ rh nor TRIM5␣ hu efficiently restricts simian immunodeficiency virus (SIV mac ) (23). Four variable regions (v1 to v4) are found in the B30.2 domains of TRIM5␣ proteins from different primates (19,21). Differences in the v1 regions of TRIM5␣ rh and TRIM5␣ hu account for the differences in anti-HIV-1 potency of these TRIM5␣ variants (15,19,24,27). Alteration of arginine 332 in the v1 region of TRIM5␣ hu to the proline residue found in TRIM5␣ rh results in a protein that can potently restrict HIV-1 and, surprisingly, SIV mac infection (24, 27). Here we investigate the specific v1 sequences in TRIM5␣ hu required for efficient antiviral activity against HIV-1 and SIV mac and provide a mechanistic explanation for the observed enhancement of restriction that results from changes in this region. MATERIALS AND METHODS Plasmids a...

No abstract