Primate genomes encode a variety of innate immune strategies to defend themselves against retroviruses. One of these, TRIM5␣, can restrict diverse retroviruses in a species-specific manner. Thus, whereas rhesus TRIM5␣ can strongly restrict HIV-1, human TRIM5␣ only has weak HIV-1 restriction. The biology of TRIM5␣ restriction suggests that it is locked in an antagonistic conflict with the proteins encoding the viral capsid. Such antagonistic interactions frequently result in rapid amino acid replacements at the proteinprotein interface, as each genetic entity vies for evolutionary dominance. By analyzing its evolutionary history, we find strong evidence for ancient positive selection in the primate TRIM5␣ gene. This selection is strikingly variable with some of the strongest selection occurring in the human lineage. This history suggests that TRIM5␣ evolution has been driven by antagonistic interactions with a wide variety of viruses and endogenous retroviruses that predate the origin of primate lentiviruses. A 13-aa ''patch'' in the SPRY protein domain bears a dense concentration of positively selected residues, potentially implicating it as an antiviral interface. By using functional studies of chimeric TRIM5␣ genes, we show that this patch is generally essential for retroviral restriction and is responsible for most of the species-specific antiretroviral restriction activity. Our study highlights the power of evolutionary analyses, in which positive selection identifies not only the age of genetic conflict but also the interaction interface where this conflict plays out.capsid ͉ human endogenous retroviruses ͉ HIV type 1 ͉ SPRY

HIV-1 is able to infect nondividing cells productively in part because the postentry viral nucleoprotein complexes are actively imported into the nucleus. In this manuscript, we identify a novel nuclear localization signal (NLS) in the viral integrase (IN) protein that is essential for virus replication in both dividing and nondividing cells. The IN NLS stimulates the efficient nuclear accumulation of viral DNA as well as virion-derived IN protein during the initial stages of infection but is dispensable for catalytic function. Because this NLS is required for infection irrespective of target cell proliferation, we suggest that interactions between uncoated viral nucleoprotein complexes and the host cell nuclear import machinery are critical for HIV-1 infection of all cells.

SUMMARY In the ubiquitin proteasome system, the E3 ligase SCF-Skp2 and its accessory protein Cks1 promote proliferation largely by inducing the degradation of the CDK inhibitor p27. Overexpression of Skp2 in human cancers correlates with poor prognosis, and deregulation of SCF-Skp2-Cks1 promotes tumorigenesis in animal models. We identified small molecule inhibitors specific to SCF-Skp2 activity using in silico screens targeted to the binding interface for p27. These compounds selectively inhibited Skp2-mediated p27 degradation by reducing p27 binding through key compound-receptor contacts. In cancer cells, the compounds induced p27 accumulation in a Skp2-dependent manner and promoted cell-type specific blocks in the G1 or G2/M phases. Designing SCF-Skp2 specific inhibitors may be a novel strategy to treat cancers dependent on the Skp2-p27 axis.

Human immunodeficiency virus type 1 (HIV-1) is a retrovirus that can cause extensive cytopathicity in T cells. However, long-term productive infection of T-cell lines has been described. Here we show that although Vpr has no effect on the initial cytopathic effect of HIV-1, viruses that contain an intact vpr gene are unable to establish a chronic infection of T cells. However, virus with a mutated vpr gene can readily establish such long-term cultures. The effect of Vpr is independent of the env gene and the nef gene. Furthermore, expression of Vpr alone affects the progression of cells in the cell cycle. These results suggest that HIV-1 has evolved a viral gene to prevent chronic infection of T cells.

CCR5 and CXCR4 are the two major coreceptors that have been identified for human immunodeficiency virus (HIV) entry. We have modified several beta-galactosidase-based HIV indicator cell lines to express CCR5 and/or CXCR4. Using these new reagents, we have been able to detect all primary isolates tested using one or both of these cell lines. However, there is large variation in the absolute viral infectivity among primary strains. Furthermore, all HIV strains are capable of causing syncytia in the indicator cells when the coreceptor is present regardless of whether they had previously been characterized as "syncytia-inducing" or "non-syncytium-inducing."

The intracellular TRIM5alpha protein successfully inhibits HIV-1 infection in rhesus monkeys, but not in humans . A few amino acids in the virus-interacting SPRY domain were found to be responsible for most of this anti-viral specificity , raising the possibility that genetic variation among humans could result in TRIM5alpha proteins with a spectrum of potencies. We found several nonsynonymous SNPs at the human TRIM5 locus, but only one of these (H43Y) was found to have a significant functional consequence. We demonstrate that H43Y impairs TRIM5alpha restriction of two distantly related retroviruses. H43Y lies in the RING domain of TRIM5alpha and may negatively affect its putative E3 ubiquitin ligase activity. This detrimental allele dates back to before the African diaspora and is found at a frequency of 43% in indigenous Central and South Americans. We suggest that relaxed constraint due to a recent period of low retroviral challenge has allowed the deleterious H43Y mutation to persist and even to expand after the bottleneck that occurred upon human migration to the New World. The unexpectedly high frequency of an impaired retroviral restriction allele among humans is likely to have a significant impact on our ability to ward off future retroviral challenges.

Humans express seven human APOBEC3 proteins, which can inhibit viruses and endogenous retroelements through cytidine deaminase activity. The seven paralogs differ in the potency of their antiviral effects, as well as in their antiviral targets. One APOBEC3, APOBEC3C, is exceptional as it has been found to only weakly block viruses and endogenous retroelements compared to other APOBEC3s. However, our positive selection analyses suggest that APOBEC3C has played a role in pathogen defense during primate evolution. Here, we describe a single nucleotide polymorphism in human APOBEC3C, a change from serine to isoleucine at position 188 (I188) that confers potent antiviral activity against HIV-1. The gain-of-function APOBEC3C SNP results in increased enzymatic activity and hypermutation of target sequences when tested in vitro, and correlates with increased dimerization of the protein. The I188 is widely distributed in human African populations, and is the ancestral primate allele, but is not found in chimpanzees or gorillas. Thus, while other hominids have lost activity of this antiviral gene, it has been maintained, or re-acquired, as a more active antiviral gene in a subset of humans. Taken together, our results suggest that APOBEC3C is in fact involved in protecting hosts from lentiviruses.

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