Destabilization of a non-enveloped virus generates a membrane transport-competent viral particle. Here we probe polyomavirus SV40 endoplasmic reticulum (ER)-to-cytosol membrane transport, a decisive infection step where destabilization initiates this non-enveloped virus for membrane penetration. We find that a member of the ER membrane protein complex (EMC) called EMC1 promotes SV40 ER membrane transport and infection. Surprisingly, EMC1 does so by using its predicted transmembrane residue D961 to bind to and stabilize the membrane-embedded partially destabilized SV40, thereby preventing premature viral disassembly. EMC1-dependent stabilization enables SV40 to engage a cytosolic extraction complex that ejects the virus into the cytosol. Thus EMC1 acts as a molecular chaperone, bracing the destabilized SV40 in a transport-competent state. Our findings reveal the novel principle that coordinated destabilization-stabilization drives membrane transport of a non-enveloped virus.

Many viral pathogens target innate sensing cascades and/or cellular transcription factors to suppress antiviral immune responses. Here, we show that the accessory viral protein U (Vpu) of HIV-1 exerts broad immunosuppressive effects by inhibiting activation of the transcription factor NF-κB. Global transcriptional profiling of infected CD4 +T cells revealed that vpu-deficient HIV-1 strains induce substantially stronger immune responses than the respective wild type viruses. Gene set enrichment analyses and cytokine arrays showed that Vpu suppresses the expression of NF-κB targets including interferons and restriction factors. Mutational analyses demonstrated that this immunosuppressive activity of Vpu is independent of its ability to counteract the restriction factor and innate sensor tetherin. However, Vpu-mediated inhibition of immune activation required an arginine residue in the cytoplasmic domain that is critical for blocking NF-κB signaling downstream of tetherin. In summary, our findings demonstrate that HIV-1 Vpu potently suppresses NF-κB-elicited antiviral immune responses at the transcriptional level.

The mammalian NF-κB p52:p52 homodimer together with its cofactor Bcl3 activates transcription of κB sites with a central G/C base pair (bp), while it is inactive toward κB sites with a central A/T bp. To understand the molecular basis for this unique property of p52, we have determined the crystal structures of recombinant human p52 protein in complex with a P-selectin(PSel)-κB DNA (5'-GGGGTGACCCC-3') (central bp is underlined) and variants changing the central bp to A/T or swapping the flanking bp. The structures reveal a nearly two-fold widened minor groove in the central region of the DNA as compared to all other currently available NF-κB-DNA complex structures, which have a central A/T bp. Microsecond molecular dynamics (MD) simulations of free DNAs and p52 bound complexes reveal that free DNAs exhibit distinct preferred conformations, and p52:p52 homodimer induces the least amount of DNA conformational changes when bound to the more transcriptionally active natural G/C-centric PSel-κB, but adopts closed conformation when bound to the mutant A/T and swap DNAs due to their narrowed minor grooves. Our binding assays further demonstrate that the fast kinetics favored by entropy is correlated with higher transcriptional activity. Overall, our studies have revealed a novel conformation for κB DNA in complex with NF-κB and pinpoint the importance of binding kinetics, dictated by DNA conformational and dynamic states, in controlling transcriptional activation for NF-κB.

The advent of sexual reproduction and the evolution of a dedicated germline in multicellular organisms are critical landmarks in eukaryotic evolution. We report an ancient family of GCNA (germ cell nuclear antigen) proteins that arose in the earliest eukaryotes, and feature a rapidly evolving intrinsically disordered region (IDR). Phylogenetic analysis reveals that GCNA proteins emerged before the major eukaryotic lineages diverged; GCNA predates the origin of a dedicated germline by a billion years. Gcna gene expression is enriched in reproductive cells across eukarya - either just prior to or during meiosis in single-celled eukaryotes, and in stem cells and germ cells of diverse multicellular animals. Studies of Gcna-mutant C. elegans and mice indicate that GCNA has functioned in reproduction for at least 600 million years. Homology to IDR-containing proteins implicated in DNA damage repair suggests that GCNA proteins may protect the genomic integrity of cells carrying a heritable genome.