Prototype foamy virus (PFV) is a member of the unconventional and nonpathogenic retroviruses. PFV causes lifelong chronic infections, which are partially attributable to a number of host cell factors that restrict viral replication. Herein, we identified human p53-induced RING-H2 protein (Pirh2) as a novel inhibitor of prototype foamy virus. Overexpression of Pirh2 decreased the replication of PFV, whereas knockdown of Pirh2 with specific siRNA increased PFV replication. Dual-luciferase assays and coimmunoprecipitation demonstrated that Pirh2 negatively influences the Tas-dependent transcriptional activation of the PFV long terminal repeat (LTR) and internal promoter (IP) by interacting with the transactivator Tas and down-regulating its expression. In addition, the viral inhibitory function of Pirh2 is N-terminal and RING domain dependent. Together, these results indicated that Pirh2 suppresses PFV replication by negatively impacting its transactivator Tas and the transcription of two viral promoters, which may contribute to the latency of PFV infection.
Prototype foamy virus (PFV) is a member of the Spumaretrovirinae subfamily of retroviruses, which maintains lifelong latent infection while being nonpathogenic to their natural hosts. Autophagy is a cell-programmed mechanism that plays a pivotal role in controlling homeostasis and defense against exotic pathogens. However, whether autophagy is the mechanism for host defense in PFV infection has not been investigated. Our results revealed that PFV infection induced the accumulation of autophagosomes and triggered complete autophagic flux in BHK-21 cells. PFV infection also altered endoplasmic reticulum (ER) homeostasis. The PERK, IRE1 and ATF6 pathways, all of which are components of the ER stress-related unfolded protein response (UPR), were activated in PFV-infected cells. In addition, accelerating autophagy suppressed PFV replication, and inhibition of autophagy promoted viral replication. Our data indicate that PFV infection can induce complete autophagy through activating the ER stress-related UPR pathway in BHK-21 cells. In turn, autophagy negatively regulates PFV replication.
HIV is the causative pathogen of AIDS, which has generated worldwide concern. Long noncoding RNAs (lncRNAs) are a rising star in virus-host cross-talk pathways; they are differentially expressed during many viral infections and are involved in multiple biological processes. Currently, lncRNA growth arrest-specific transcript 5 (GAS5) is known to be downregulated during HIV-1 infection. However, the functions and mechanisms of GAS5 in HIV-1 infection remain largely unknown. In this report, it was found for the first time that GAS5 could inhibit HIV-1 replication. Interestingly, using bioinformatics analyses (with Genomica and starBase.v2.0), GAS5 was found to potentially interact with miR-873. It was further verified that GAS5 could suppress miR-873. Moreover, miR-873 could promote HIV-1 replication. Together, these results not only suggest that GAS5 may inhibit HIV-1 replication through interaction with miR-873 but the results may also provide novel biomarkers for antiviral drugs or potential targets for future therapeutics for HIV/AIDS.
Human foamy virus (HFV) is a complex and unique retrovirus with the longest genomes among retroviruses that are used as vectors for gene therapy. Long non-coding RNAs (lncRNAs) are regarded as key regulators that are involved in diverse biological processes during viral infection. However, the role of lncRNAs in HFV infection remains unknown. In this study, we utilized next-generation sequencing to first characterize lncRNAs in 293T cells after HFV infection, evaluating length distribution, exon number distribution, volcano picture, and lncRNA class distribution. We identified 11,336 lncRNAs (4,729 upregulated lncRNAs and 6,588 downregulated lncRNAs) and 61,367 mRNAs (30,133 upregulated mRNAs and 31,220 downregulated mRNAs), which were differentially expressed in the HFV-infected 293T cells. Subsequently, six differentially expressed lncRNAs characterized using RNA-seq were confirmed by quantitative real-time polymerase chain reaction assays. Interestingly, Gene Ontology (GO)/Gene Ontology Tree Machine (GOTM) and Kyoto Encyclopedia of Gene and Genomes (KEGG) pathway analyses indicated that positive regulation of interleukin 8 (IL8) production and cytokine-cytokine receptor interaction might be involved in the functional enrichment of lncRNAs. Moreover, cis-acting and trans-acting regulatory networks show that NR_028036 may target the fas gene in a cis-acting manner and that ENST00000354838 may target the IL18 gene in a trans-acting manner. Overall, these results not only provide novel insights into the relationship between HFV and lncRNAs in the host response to infection but also have implications for the future wider application of HFV as a vector.
RNA interference has shown great potential for the treatment of HIV-1. Vectors derived from prototype foamy viruses (PFVs) with a nonpathogenic nature are very promising gene transfer vehicles in anti-HIV gene therapy. In this article, three short hairpin RNAs (shRNAs) targeting the conserved regions of the HIV-1NL4-3 5' long terminal repeat (LTR) were first designed. We then constructed novel recombinant PFV vector plasmids, p▵Φ-H1-shRNAs, expressing these shRNAs under the control of the H1 RNA promoter. To detect the efficacy of these ▵Φ-H1-shRNAs for the inhibition of HIV-1 replication, we performed a dual-luciferase reporter assay, RT-qPCR, ELISA, western blotting, and a lactate dehydrogenase (LDH) assay by transient transfection in 293T cells. The results suggest that these novel shRNAs driven by PFV vectors inhibit HIV-1 replication efficiently without cytotoxicity, with shRNA3 being the most effective. In addition, we analyzed the shRNA target sites in the 5' LTR of HIV-1 strains other than HIV-1NL4-3 and found that these shRNAs may possibly inhibit other HIV-1 strains.
Prototype foamy virus (PFV) is a member of the oldest family of retroviruses and maintains lifelong latent infection in the host. The lifelong latent infection of PFV may be maintained by the restriction factors of viral replication in the host. However, the mechanisms involved in PFV latent infection are poorly understood. Here, we found that TBC1D16, a TBC domain-containing protein, is significantly down-regulated after PFV infection. Tre2/Bub2/Cdc16 (TBC) domain-containing proteins function as Rab GTPase-activating proteins (GAPs) and are participates in the progression of some diseases and many signaling pathways. However, whether TBC proteins are involved in PFV replication has not been determined. Here, we found that TBC1D16 is a novel antiviral protein that targets Rab5C to suppress PFV replication. Overexpression TBC1D16 inhibited the transcription and expression of Tas and Gag, and silencing TBC1D16 enhanced the PFV replication. Moreover, the highly conserved amino acid residues R494 and Q531 in the TBC domain of TBC1D16 were essential for inhibiting PFV replication. We also found that TBC1D16 promoted the production of PFV-induced IFN-β and the transcription of downstream genes. These results suggest that TBC1D16 might be the first identified TBC proteins that inhibited PFV replication and the mechanism by which TBC1D16 inhibited PFV replication could provide new insights for PFV latency.
Abstract Background Prototype foamy virus (PFV) is nonpathogenic complex retroviruses that express a transcriptional transactivator Tas, which is essential for the activity of viral long terminal repeat (LTR) promoter and internal promoter (IP). Tripartite motif-containing protein 28 (Trim28) is well known as a scaffold protein normally enriched in gene promoter region to repress transcription. We sought to determine if whether Trim28 could be enriched in PFV promoter region to participate the establishment of PFV latency infection. Results In this study, we show that Trim28 restricts Tas-dependent transactivation activity of PFV promoter and negatively regulates PFV replication. Trim28 was found to be enriched in LTR instead of IP promoter regions of PFV genome and contribute to the maintenance of histone H3K9me3 marks on the LTR promoter. Furthermore, Trim28 interacts with Tas and colocalizes with Tas in the nucleus. Besides, we found that Trim28, an E3 ubiquitin ligase, binds directly to and promotes Tas for ubiquitination and degradation. And the RBCC domain of Trim28 is required for the ubiquitination and degradation of Tas. Conclusions Collectively, our findings not only identify a host factor Trim28 negatively inhibits PFV replication by acting as transcriptional restriction factor enriched in viral LTR promoter through modulating H3K9me3 mark here, but also reveal that Trim28 mediated ubiquitin proteasome degradation of Tas as a mechanism underlying Trim28 restricts Tas-dependent transcription activity of PFV promoter and PFV replication. These findings provide new insights into the process of PFV latency establishment. Graphical Abstract
<b><i>Background:</i></b> Prototype foamy virus (PFV) is a complex and unique retrovirus with the longest genome among the retroviruses and is used as a vector for gene therapies. The viral Tas protein transactivates the viral long terminal repeat promoter and is required for viral replication. We have utilized RNA sequencing to identify and characterize the long-noncoding RNA NONHSAG000101 (lnc-NONH), which markedly increases in PFV-infected cells. However, little is known about the function of lnc-NONH. <b><i>Objectives:</i></b> We aim to explore the role of lnc-NONH during PFV infection. <b><i>Methods:</i></b> To assess the lnc-NONH role during PFV infection, the siRNAs were used to silence the lnc-NONH expression. The microRNA (miRNA) mimic and inhibitor were employed to explore the function of lnc-NONH-related miRNA miR-34c-5p. Quantitative real-time polymerase chain reaction assay and Western blotting were applied to measure the mRNA and protein levels of PFV transactivator Tas. Luciferase assay was used to determine the transcriptional activity of the PFV unique internal promoter (IP). <b><i>Results:</i></b> lnc-NONH promotes the expression of PFV Tas and miR-34c-5p. The interaction between lnc-NONH and miR-34c-5p enhances the transcriptional activity of the PFV IP. <b><i>Conclusions:</i></b> In the current study, we report a novel mechanism for the lnc-NONH-mediated upregulation of Tas expression. Our findings contribute to the understanding of regulatory network of Tas expression and PFV replication.
Prototype foamy virus (PFV) is an ancient retrovirus that infects humans with persistent latent infections and non-pathogenic consequences. Lifelong latent PFV infections can be caused by restrictive factors in the host. However, the molecular mechanisms underlying host cell regulation during PFV infection are not fully understood. The aim of the study was to investigate whether a zinc finger protein (ZFP), ZNF219, as a transcription factor, can regulate the transcriptional activity of the viral promoter. Here, using transcriptome sequencing, we found that ZNF219, is downregulated in PFV infected cells and that ZNF219 suppresses viral replication by targeting the viral 5'LTR promoter region to repress its transcription. We also found that PFV infection induced abnormal expression of miRNAs targeting the ZNF219-3'UTR to downregulate ZNF219 expression. These findings indicated that ZNF219 may be a potent antiviral factor for suppressing PFV infection, and may shed light on the mechanism of virus-host interactions.
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