TRIM5α recruits HDAC1 to p50 and Sp1 and promotes H3K9 deacetylation at the HIV-1 LTR.

Tripartite motif-containing protein 5α (TRIM5α) is generally known to block the postentry events of HIV-1. Here, we report an uncharacterized role for TRIM5α in the maintenance of viral latency. Knockdown of TRIM5α potentiates the transcription of HIV-1 in multiple latency models, which is reversed by shRNA-resistant TRIM5α. TRIM5α suppresses TNFα-activated HIV-1 LTR-driven as well as NF-κB- and Sp1-driven gene expression, with the RING and B-box 2 domains being the essential determinants. Mechanistically, TRIM5α binds to and enhances the recruitment of histone deacetylase 1 (HDAC1) to NF-κB p50 and Sp1. ChIP‒qPCR analyses further reveal that the association of TRIM5α with HIV-1 LTR induces HDAC1 recruitment and local H3K9 deacetylation. Conserved suppression effects of TRIM5α orthologs from multiple species on both HIV-1 and endo-retroelement HERV-K LTR activities have also been demonstrated. These findings provide new insights into the molecular mechanisms by which proviral latency is initially established and activatable proviruses are resilenced by histone deacetylase recruitment.

TRIM28 negatively regulates the RLR signaling pathway by targeting MAVS for degradation via K48-linked polyubiquitination.

Mitochondrial antiviral signaling (MAVS) protein is a core signaling adapter in the retinoid acid-inducible gene-I-like receptor (RLR) signaling pathway that recruits downstream signaling factors, ultimately leading to the activation of type Ⅰ interferons. However, the mechanisms that modulate the RLR signaling pathway by manipulating MAVS are not fully understood. Previous studies suggested that tripartite motif 28 (TRIM28) participates in regulating innate immune signaling pathways by inhibiting the expression of immune-related genes at the transcriptional level. In this study, we characterized TRIM28 as a negative regulator of the RLR signaling pathway in a MAVS-dependent manner. Overexpression of TRIM28 inhibited the MAVS-induced production of type Ⅰ interferons and proinflammatory cytokines, while knocking down TRIM28 exerted the opposite effect. Mechanistically, TRIM28 targeted MAVS for proteasome-mediated degradation via K48-linked polyubiquitination. The RING domain of TRIM28, especially the cysteine residues at positions 65 and 68, was critical for the suppressive effect of TRIM28 on MAVS-mediated RLR signaling, while each of the C-terminal domains of TRIM28 contributed to its interaction with MAVS. Further investigation revealed that TRIM28 transferred ubiquitin chains to the K7, K10, K371, K420, and K500 residues of MAVS. Together, our results reveal a previously uncharacterized mechanism involving TRIM28 in fine-tuning innate immune responses and provide new insights into the mechanisms by which MAVS is regulated, which contribute to the understanding of the molecular mechanisms underlying immune homeostasis maintenance.

Papain-like protease of SARS-CoV-2 inhibits RLR signaling in a deubiquitination-dependent and deubiquitination-independent manner.

The newly emerged severe acute respiratory syndrome (SARS) coronavirus-2 (SARS-CoV-2) can result in dysregulated interferon (IFN) responses that contribute to disease severity. The papain-like protease of SARS-CoV-2 (SCoV2-PLpro) has been previously reported to attenuate IFN responses, but the underlying mechanism is not fully understood. In this study, we found that SCoV2-PLpro potently suppressed IFN production and signaling induced by Sendai virus as well as RIG-I-like receptor (RLR) signaling pathway components, including RIG-I, MAVS, TBK1, TRAF3, TRAF6, and IRF3. SCoV2-PLpro exhibited different specificity and efficiency than SARS-CoV PLpro, with the former exerting a greater inhibitory effect on the RIG-I- and TRAF3-mediated IFN response but a weaker effect on the MAVS-mediated IFN response. Furthermore, we showed that SCoV2-PLpro significantly reduced K63-ubiquitination of RIG-I, MAVS, TBK1, TRAF3, TRAF6, and IRF3 and K48-ubiquitination of IκBα, which are known critical for the innate immune signal transduction. The deubiquitinating (DUB) activity of SCoV2-PLpro required a catalytic residue cysteine 111 (C111) but not the UBL domain. Notably, by utilizing the DUB-defective C111 mutant, we demonstrated that SCoV2-PLpro targeted RLR signaling pathway regulators via deubiquitination-dependent and -independent mechanisms, with the inhibitory activities of RIG-I and TBK1 correlating with DUB function, whereas the antagonism effects on MAVS, TRAF3, TRAF6, and IRF3 independent on DUB activity. Overall, our results reveal that SCoV2-PLpro evolves differential IFN antagonism activity from SCoV1-PLpro and it targets multiple key RLR signaling pathway components via various mechanisms, providing insights into SARS-CoV-2 pathogenesis and clues for developing antiviral therapies for COVID-19.