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Intracellular innate immunity involves co-evolved antiviral restriction factors that specifically inhibit infecting viruses. Studying these restrictions has increased our understanding of viral replication, host-pathogen interactions, and pathogenesis, and represent potential targets for novel antiviral therapies. Lentiviral restriction 2 (Lv2) was identified as an unmapped early-phase restriction of HIV-2 and later shown to also restrict HIV-1 and simian immunodeficiency virus. The viral determinants of Lv2 susceptibility have been mapped to the envelope and capsid proteins in both HIV-1 and HIV-2, and also viral protein R (Vpr) in HIV-1, and appears dependent on cellular entry mechanism. A genome-wide screen identified several likely contributing host factors including members of the polymerase-associated factor 1 (PAF1) and human silencing hub (HUSH) complexes, and the newly characterized regulation of nuclear pre-mRNA domain containing 2 (RPRD2). Subsequently, RPRD2 (or RNA-associated early-stage antiviral factor) has been shown to be upregulated upon T cell activation, is highly expressed in myeloid cells, binds viral reverse transcripts, and potently restricts HIV-1 infection. RPRD2 is also bound by HIV-1 Vpr and targeted for degradation by the proteasome upon reverse transcription, suggesting RPRD2 impedes reverse transcription and Vpr targeting overcomes this block. RPRD2 is mainly localized to the nucleus and binds RNA, DNA, and DNA:RNA hybrids. More recently, RPRD2 has been shown to negatively regulate genome-wide transcription and interact with the HUSH and PAF1 complexes which repress HIV transcription and are implicated in maintenance of HIV latency. In this review, we examine Lv2 restriction and the antiviral role of RPRD2 and consider potential mechanism(s) of action.

Human immunodeficiency virus (HIV) relies upon a broad array of host factors in order to replicate and evade the host antiviral response. Cleavage and polyadenylation specificity factor 6 (CPSF6) is one such host factor that is recruited by incoming HIV-1 cores to regulate trafficking, nuclear import, uncoating, and integration site selection. Despite these well-described roles, the impact of CPSF6 perturbation on HIV-1 infectivity varies considerably by cell type. Here, we report that CPSF6 knock-out in primary CD4+ T cells leads to increased permissivity to HIV-1 infection due to broad transcriptional reprogramming. Knock-out of CPSF6 results in widespread differential gene expression, including downregulation of genes involved in the innate immune response and enhanced expression of the HIV-1 co-receptors. Accordingly, these cells are less responsive to interferon and express lower levels of antiretroviral restriction factors, including TRIM5α. These transcriptional changes are linked to global shortening of mRNA 3' untranslated regions (UTRs) through changes in alternative polyadenylation (APA), which are triggered by disruption of the CPSF6-containing Cleavage Factor Im (CFIm) complex. Furthermore, we find that recruitment of CPSF6 by HIV-1 cores is sufficient to perturb CPSF6 function, leading to 3' UTR shortening and subsequent transcriptional rewiring. These results suggest a model in which HIV-1 transcriptionally reprograms target cells through recruitment of CPSF6 to incoming cores to circumvent the antiviral response and enhance permissivity to infection.

N -methyladenosine (m A) is the most prevalent internal modification of cellular and viral RNA and is critical to the regulation of its localization, stability, and translation. Previous studies on the role of m A during HIV-1 replication have produced conflicting results. Since m A function can vary dramatically by cell type and state, here we aimed to clarify the role of the m A machinery during HIV-1 replication in primary CD4+ T cells. Using CRISPR-Cas9 we targeted 46 cellular genes implicated in m A or 5-methylcytosine (m C) regulation and measured subsequent HIV-1 replication in primary CD4+ T cells. Only knockout of the m A writer complex auxiliary proteins VIRMA and WTAP, and the m A reader YTHDF2 were validated as significantly decreasing HIV-1 replication. In contrast, knockout of METTL3 or METTL14, which form the catalytic core of the writer complex, resulted in only marginal changes in HIV-1 infection, despite significant decreases in total cellular m A levels. Chemical inhibition of METTL3 led to a dose-dependent decrease in HIV-1 infection, coupled with an increase in protein levels of METTL3 and other writer complex members. Expression of writer proteins was also co-dependent, revealing complex regulatory feedback mechanisms. Overall, these results clarify the role of epitranscriptomic machinery during HIV-1 replication in primary CD4+ T cells and suggest regulation by auxiliary members of the m A writer complex is more influential than the function of the catalytic core itself on HIV-1 infection in primary CD4+ T cells.

Nonsense-mediated decay (NMD) is a translation-dependent RNA quality control mechanism that occurs in the cytoplasm. However, it is unknown how NMD regulates the stability of RNAs translated at the Endoplasmic Reticulum (ER). Here, we identify a localized NMD pathway dedicated to ER-translated mRNAs. We previously identified NBAS, a component of the Syntaxin 18 complex involved in Golgi-to-ER trafficking, as a novel NMD factor. Here, we show that NBAS fulfils an independent function in NMD. This ER-NMD pathway requires the interaction of NBAS with the core NMD factor UPF1, which is partially localized at the ER in the proximity of the translocon. NBAS and UPF1 co-regulate the stability of ER-associated transcripts, in particular those associated with the cellular stress response. We propose a model where NBAS recruits UPF1 to the membrane of the ER and activates an ER-dedicated NMD pathway, thus providing an ER protective function by ensuring quality control of ER-translated mRNAs. NBAS is an NMD factor that localizes to the membrane of the endoplasmic reticulum (ER) NBAS has dual, independent, roles in Golgi-to-ER retrograde transport and in ER-NMD NBAS recruits the core NMD factor UPF1 to the membrane of the ER The ER-NMD pathway targets for degradation mRNAs that are translated at the ER