The roles of nuclear orphan receptor NR2F6 in anti-viral innate immunity.

Proper transcription regulation by key transcription factors, such as IRF3, is critical for anti-viral defense. Dynamics of enhancer activity play important roles in many biological processes, and epigenomic analysis is used to determine the involved enhancers and transcription factors. To determine new transcription factors in anti-DNA-virus response, we have performed H3K27ac ChIP-Seq and identified three transcription factors, NR2F6, MEF2D and MAFF, in promoting HSV-1 replication. NR2F6 promotes HSV-1 replication and gene expression in vitro and in vivo, but not dependent on cGAS/STING pathway. NR2F6 binds to the promoter of MAP3K5 and activates AP-1/c-Jun pathway, which is critical for DNA virus replication. On the other hand, NR2F6 is transcriptionally repressed by c-Jun and forms a negative feedback loop. Meanwhile, cGAS/STING innate immunity signaling represses NR2F6 through STAT3. Taken together, we have identified new transcription factors and revealed the underlying mechanisms involved in the network between DNA viruses and host cells.

Histone demethylase LSD1 promotes RIG-I poly-ubiquitination and anti-viral gene expression.

Under RNA virus infection, retinoic acid-inducible gene I (RIG-I) in host cells recognizes viral RNA and activates the expression of type I IFN. To investigate the roles of protein methyltransferases and demethylases in RIG-I antiviral signaling pathway, we screened all the known related enzymes with a siRNA library and identified LSD1 as a positive regulator for RIG-I signaling. Exogenous expression of LSD1 enhances RIG-I signaling activated by virus stimulation, whereas its deficiency restricts it. LSD1 interacts with RIG-I, promotes its K63-linked polyubiquitination and interaction with VISA/MAVS. Interestingly, LSD1 exerts its function in antiviral response not dependent on its demethylase activity but through enhancing the interaction between RIG-I with E3 ligases, especially TRIM25. Furthermore, we provide in vivo evidence that LSD1 increases antiviral gene expression and inhibits viral replication. Taken together, our findings demonstrate that LSD1 is a positive regulator of signaling pathway triggered by RNA-virus through mediating RIG-I polyubiquitination.

Removal of epigenetic repressive mark on inflammatory genes in fat liver.

Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disease worldwide. The detailed epigenomic changes during fat accumulation in liver are not clear yet. Here, we performed ChIP-Seq analysis in the liver tissues of high-fat diet and regular chow diet mice and investigated the dynamic landscapes of H3K27ac and H3K9me3 marks on chromatin. We find that the activated typical enhancers marked with H3K27ac are enriched on lipid metabolic pathways in fat liver; however, super enhancers do not change much. The regions covered with H3K9me3 repressive mark seem to undergo great changes, and its peak number and intensity both decrease in fat liver. The enhancers located in lost H3K9me3 regions are enriched in lipid metabolism and inflammatory pathways; and motif analysis shows that they are potential targets for transcription factors involved in metabolic and inflammatory processes. Our study has revealed that H3K9me3 may play an important role during the pathogenesis of NAFLD through regulating the accessibility of enhancers.

Deficiency of Histone Methyltransferase SET Domain-Containing 2 in Liver Leads to Abnormal Lipid Metabolism and HCC.

BACKGROUND AND AIMS: Trimethylation of Lys36 on histone 3 (H3K36me3) catalyzed by histone methyltransferase SET domain-containing 2 (SETD2) is one of the most conserved epigenetic marks from yeast to mammals. SETD2 is frequently mutated in multiple cancers and acts as a tumor suppressor. APPROACH AND RESULTS: Here, using a liver-specific Setd2 depletion model, we found that Setd2 deficiency is sufficient to trigger spontaneous HCC. Meanwhile, Setd2 depletion significantly increased tumor and tumor size of a diethylnitrosamine-induced HCC model. The mechanistic study showed that Setd2 suppresses HCC not only through modulating DNA damage response, but also by regulating lipid metabolism in the liver. Setd2 deficiency down-regulated H3K36me3 enrichment and expression of cholesterol efflux genes and caused lipid accumulation. High-fat diet enhanced lipid accumulation and promoted the development of HCC in Setd2-deficient mice. Chromatin immunoprecipitation sequencing analysis further revealed that Setd2 depletion induced c-Jun/activator protein 1 (AP-1) activation in the liver, which was trigged by accumulated lipid. c-Jun acts as an oncogene in HCC and functions through inhibiting p53 in Setd2-deficient cells. CONCLUSIONS: We revealed the roles of Setd2 in HCC and the underlying mechanisms in regulating cholesterol homeostasis and c-Jun/AP-1 signaling.

SETD8 involved in the progression of inflammatory bowel disease via epigenetically regulating p62 expression.

BACKGROUND AND AIM: Epigenetic modification is an important part of the pathogenesis of inflammatory bowel disease (IBD). Some studies proved that p62 was involved in inflammatory response and upregulated in IBD patients, and histone modification plays an important role in regulating p62 expression. SETD8, a histone H4K20 methyltransferase, has been reported downregulated in some inflammatory diseases. Here, we investigated the role of SETD8 in the development of IBD and its underlying mechanisms. METHODS: An inflammatory cell model was established to elucidate whether SETD8 involved in inflammatory response in macrophages. Three percent dextran sodium sulfate-induced colitis murine model injection with SETD8 inhibitor was used in our study to investigate whether SETD8 inhibition can affect the progress of IBD. The expression of SETD8 and p62 was measured by qRT-PCR and western blot. The mRNA level of inflammatory cytokines was analyzed by qRT-PCR. In addition, chromatin immunoprecipitation-PCR was performed to identify the mechanism by which SETD8 regulates p62. RESULTS: SETD8 expression obviously decreased in vitro, in vivo models and in IBD patients. In lipopolysaccharide-activated RAW264.7 cells, knockdown of SETD8 significantly increased the mRNA expression of inducible nitric oxide synthase, cyclooxygenase-2, TNF-α, IL-6, IL-1ß, and MCP-1. Based on the dataset, we verified that p62 was a target gene of SETD8 and chromatin immunoprecipitation-PCR assay identified that silence of SETD8 distinctly decreases the H4K20me1 enrichment in the promoter of p62. Moreover, silencing of p62 partly reverses the SETD8 inhibition-mediated pro-inflammatory effect in vitro. Finally, SETD8 pharmacological inhibitor (UNC0379) aggravated the disease progression in dextran sodium sulfate-induced murine colitis. CONCLUSION: Our findings elucidate an epigenetic mechanism by which SETD8 regulates the p62 expression and restrains the inflammatory response in colitis. Our result suggests that targeting SETD8 may be a promising therapy for IBD.

Histone demethylase KDM3A is required for enhancer activation of hippo target genes in colorectal cancer.

Hippo pathway is involved in tumorigenesis, and its regulation in cytosol has been extensively studied, but its regulatory mechanisms in the nuclear are not clear. In the current study, using a FBS-inducing model following serum starvation, we identified KDM3A, a demethylase of histone H3K9me1/2, as a positive regulator for hippo target genes. KDM3A promotes gene expression through two mechanisms, one is to upregulate YAP1 expression, and the other is to facilitate H3K27ac on the enhancers of hippo target genes. H3K27ac upregulation is more relevant with gene activation, but not H3K4me3; and KDM3A depletion caused H3K9me2 upregulation mainly on TEAD1-binding enhancers rather than gene bodies, further resulting in H3K27ac decrease, less TEAD1 binding on enhancers and impaired transcription. Moreover, KDM3A is associated with p300 and required for p300 recruitment to enhancers. KDM3A deficiency delayed cancer cell growth and migration, which was rescued by YAP1 expression. KDM3A expression is correlated with YAP1 and hippo target genes in colorectal cancer patient tissues, and may serve as a potential prognosis mark. Taken together, our study reveals novel mechanisms for hippo signaling and enhancer activation, which is critical for tumorigenesis of colorectal cancer.

Characterization of WDR20: A new regulator of the ERAD machinery.

ERAD is an important process of protein quality control that eliminates misfolded or unassembled proteins from ER. Before undergoing proteasome degradation, the misfolded proteins are dislocated from ER membrane into cytosol, which requires the AAA ATPase p97/VCP and its cofactor, the NPL4-UFD1 dimer. Here, we performed a CRISPR-based screen and identify many candidates for ERAD regulation. We further confirmed four proteins, FBOX2, TRIM6, UFL1 and WDR20, are novel regulators for ERAD. Then the molecular mechanism for WDR20 in ERAD is further characterized. Depletion of WDR20 inhibits the degradation of TCRα, a typical ERAD substrate, while WDR20 overexpression reduces TCRα protein level. WDR20 associates with TCRα and central regulators of the ERAD system, p97, GP78 and HRD1. A portion of WDR20 localizes to the ER-containing microsomal membrane. WDR20 expression increases TCRα ubiquitination, and HRD1 E3 ligase is essential for the process. WDR20 seems to serve as an adaptor protein to mediate the interaction between p97 and TCRα. Our study provides novel candidates and reveals an unexpected role of WDR20 in ERAD regulation.

TTLL12 Inhibits the Activation of Cellular Antiviral Signaling through Interaction with VISA/MAVS.

Upon virus infection, host cells use retinoic-acid-inducible geneI I (RIG-I)-like receptors to recognize viral RNA and activate type I IFN expression. To investigate the role of protein methylation in the antiviral signaling pathway, we screened all the SET domain-containing proteins and identified TTLL12 as a negative regulator of RIG-I signaling. TTLL12 contains SET and TTL domains, which are predicted to have lysine methyltransferase and tubulin tyrosine ligase activities, respectively. Exogenous expression of TTLL12 represses IFN-ß expression induced by Sendai virus. TTLL12 deficiency by RNA interference and CRISPR-gRNA techniques increases the induced IFN-ß expression and inhibits virus replication in the cell. The global gene expression profiling indicated that TTLL12 specifically inhibits the expression of the downstream genes of innate immunity pathways. Cell fractionation and fluorescent staining indicated that TTLL12 is localized in the cytosol. The mutagenesis study suggested that TTLL12's ability to repress the RIG-I pathway is probably not dependent on protein modifications. Instead, TTLL12 directly interacts with virus-induced signaling adaptor (VISA), TBK1, and IKKε, and inhibits the interactions of VISA with other signaling molecules. Taken together, our findings demonstrate TTLL12 as a negative regulator of RNA-virus-induced type I IFN expression by inhibiting the interaction of VISA with other proteins.

SPOP-containing complex regulates SETD2 stability and H3K36me3-coupled alternative splicing.

Trimethylation of histone H3K36 is a chromatin mark associated with active gene expression, which has been implicated in coupling transcription with mRNA splicing and DNA damage response. SETD2 is a major H3K36 trimethyltransferase, which has been implicated as a tumor suppressor in mammals. Here, we report the regulation of SETD2 protein stability by the proteasome system, and the identification of SPOP, a key subunit of the CUL3 ubiquitin E3 ligase complex, as a SETD2-interacting protein. We demonstrate that SPOP is critically involved in SETD2 stability control and that the SPOP/CUL3 complex is responsible for SETD2 polyubiquitination both in vivo and in vitro ChIP-Seq analysis and biochemical experiments demonstrate that modulation of SPOP expression confers differential H3K36me3 on SETD2 target genes, and induce H3K36me3-coupled alternative splicing events. Together, these findings establish a functional connection between oncogenic SPOP and tumor suppressive SETD2 in the dynamic regulation of gene expression on chromatin.

Inhibition of H3K4 demethylation induces autophagy in cancer cell lines.

Epigenetic factors and related small molecules have emerged to be strongly involved in autophagy process. Here we report that 2-PCPA and GSK-LSD1, two inhibitors of histone H3K4 demethylase KDM1A/LSD1, induce autophagy in multiple mammalian cell lines. The two small molecules induce accumulation of LC3II, formation of autophagosome and autolysosome, and SQSTM1/p62 degradation. 2-PCPA treatment inhibits cell proliferation through cell cycle arrest but does not inducing cell death. Exogenous expression of KDM1A/LSD1 impaired the autophagic phenotypes triggered by 2-PCPA. The autophagy induced by 2-PCPA requires LC3-II processing machinery. But depletion of BECN1 and ULK1 with siRNA did not affect the LC3-II accumulation triggered by 2-PCPA. 2-PCPA treatment induces the change of global gene expression program, including a series of autophagy-related genes, such as SQSTM1/p62. Taken together, our data indicate that KDM1A/LSD1 inhibitors induce autophagy through affecting the expression of autophagy-related genes and in a BECN1-independent manner.

The EZH1-SUZ12 complex positively regulates the transcription of NF-κB target genes through interaction with UXT.

Unlike other members of the polycomb group protein family, EZH1 has been shown to positively associate with active transcription on a genome-wide scale. However, the underlying mechanism for this behavior still remains elusive. Here, we report that EZH1 physically interacts with UXT, a small chaperon-like transcription co-activator. UXT specifically interacts with EZH1 and SUZ12, but not EED. Similar to upon knockdown of UXT, knockdown of EZH1 or SUZ12 through RNA interference in the cell impairs the transcriptional activation of nuclear factor (NF)-κB target genes induced by TNFα. EZH1 deficiency also increases TNFα-induced cell death. Interestingly, chromatin immunoprecipitation and the following next-generation sequencing analysis show that H3K27 mono-, di- and tri-methylation on NF-κB target genes are not affected in EZH1- or UXT-deficient cells. EZH1 also does not affect the translocation of the p65 subunit of NF-κB (also known as RELA) from the cytosol to the nucleus. Instead, EZH1 and SUZ12 regulate the recruitment of p65 and RNA Pol II to target genes. Taken together, our study shows that EZH1 and SUZ12 act as positive regulators for NF-κB signaling and demonstrates that EZH1, SUZ12 and UXT work synergistically to regulate pathway activation in the nucleus.

WDR82 Negatively Regulates Cellular Antiviral Response by Mediating TRAF3 Polyubiquitination in Multiple Cell Lines.

Upon virus infection, retinoic acid-inducible gene I-like receptors in host cells recognize viral RNA and activate type I IFN expression. Previously, we identified WD repeat domain (WDR) 5 as one positive regulator for pathway activation. In this study, we report that WDR82, a homolog protein of WDR5, acts opposite to WDR5 and inhibits the activation of the retinoic acid-inducible gene I signaling pathway. WDR82 overexpression inhibits virus-triggered pathway activation, whereas its knockdown enhances induced IFN-ß expression. WDR82 is localized on the mitochondria, and its first N-terminal WD40 domain is critical for localization. WDR82 interacts with TNFR-associated factor (TRAF) 3, and its overexpression promotes K48-linked, but not K63-linked, polyubiquitination on TRAF3. Furthermore, WDR82 knockdown inhibits viral replication in the cell, whereas its overexpression has the opposite effect. Interestingly, WDR82 regulates Sendai virus-induced IFNB1 expression in a cell type-specific manner. Taken together, our findings demonstrate that WDR82 is a negative regulator of virus-triggered type I IFNs pathway through mediating TRAF3 polyubiquitination status and stability on mitochondria.

Histone H3K4 methyltransferase Mll1 regulates protein glycosylation and tunicamycin-induced apoptosis through transcriptional regulation.

Disrupting protein glycosylation induces ER (endoplasmic reticulum) stress, resulting in the activation of UPR (unfolded protein response) pathways. A key function of the UPR is to restore ER homeostasis, but prolonged or unsolved ER stress can lead to apoptosis. MLL1 (Mixed Lineage Leukemia 1, also named ALL-1 or HRX), a histone H3K4 methyltransferase in mammals, plays important roles in leukemogenesis, transcriptional regulation, cell cycle and development. Here, we find that Mll1 deficiency enhances UPR and apoptosis induced by the glycosylation inhibitor TM (tunicamycin). The abnormal regulation of the UPR appears to be caused by a defect in protein glycosylation. Furthermore, Mll1 directly binds to the promoters of H6pd, Galnt12 and Ugp2, which regulates H3K4 trimethylation and the subsequent expression of these genes. The knockdown of H6pd, Galnt12 or Ugp2 enhances TM-induced apoptosis in Mll1(+/+)MEF cells, whereas the ectopic expression of these proteins inhibits TM-induced apoptosis in Mll1(-/-) MEF cells. Together, our data suggest that the maturation of glycoproteins in the ER is subject to regulation at the epigenetic level by a histone methyltransferase whose abnormality can lead to cancer and developmental defects.

MLL1, a H3K4 methyltransferase, regulates the TNFα-stimulated activation of genes downstream of NF-κB.

Genes of the mixed lineage leukemia (MLL) family regulate transcription by methylating histone H3K4. Six members of the MLL family exist in humans, including SETD1A, SETD1B and MLL1-MLL4. Each of them plays non-redundant roles in development and disease genesis. MLL1 regulates the cell cycle and the oscillation of circadian gene expression. Its fusion proteins are involved in leukemogenesis. Here, we studied the role of MLL1 in innate immunity and found it selectively regulates the activation of genes downstream of NF-κB mediated by tumor necrosis factor (TNFα) and lipopolysaccharide (LPS). Real-time PCR and genome-wide gene expression profile analysis proved that the deficiency of MLL1 reduced the expression of a group of genes downstream of nuclear factor κB (NF-κB). However, the activation of NF-κB itself was not affected. The MLL1 complex is found both in the nucleus and cytoplasm and is associated with NF-κB. CHIP assays proved that the translocation of MLL1 to chromatin was dependent on NF-κB. Our results suggest that MLL1 is recruited to its target genes by activated NF-κB and regulates their transcription.

Design, Synthesis and Evaluation of A Novel Teoptinib Derivative as an Effective Anti-Hepatocellular Carcinoma Agent.

BACKGROUND & PURPOSE: Hepatocellular Carcinoma (HCC) is a type of liver cancer known for its poor prognosis and high mortality. Teoptinib is a highly selective MET inhibitor that has been used in the treatment of liver cancer. Although good progress has been made in clinical treatment, further improvement is still needed. In this study, a series of novel Teoptinib derivatives were synthesized and evaluated as anti-cancer agents for the treatment of liver cancer, and an oral nanodrug delivery system was also explored. METHODS: A series of novel Teoptinib derivatives were synthesized, and an oral nanodrug delivery system was also explored. HPLC, high-resolution mass spectrometer and NMR were used to determine the structure and molecular formula of the synthesized compounds. Zeta potential assay was used to access the particle size distribution and zeta potential of the nanoparticles. MTT assay, cell colony formation assay, cell apoptosis inhibition assay, cell scratch assay, and the MHCC-97H xenograft model of nude mice assay were used to evaluate the in vitro and in vivo anti-tumor activity of the synthesized compounds. RESULTS: Compound (R)-10 showed the best antitumor activity with 0.010 µM of the IC50 value against MHCC-97H, a human liver cancer cell line with high c-Met expression. The MHCC-97H xenograft model of nude mice assay showed that nano-prodrug of compound (R)-10 exhibited good in vivo activity with 87.67% of the TGI at the dosage of 8 mg/kg. CONCLUSION: We designed and synthesized a series of c-Met inhibitors containing different side chains and chiral centers as anti-liver cancer agents. Among them, compound (R)-10 shows a promising effect as a lead molecule for further study in the treatment of liver cancer. The successful incorporation of (R)-10 into a novel oral nanodrug delivery system highlights the importance of effective drug delivery systems for enhanced therapeutic efficacy.

Cooperation of MLL1 and Jun in controlling H3K4me3 on enhancers in colorectal cancer.

BACKGROUND: Enhancer dysregulation is one of the important features for cancer cells. Enhancers enriched with H3K4me3 have been implicated to play important roles in cancer. However, their detailed features and regulatory mechanisms have not been well characterized. RESULTS: Here, we profile the landscape of H3K4me3-enriched enhancers (m3Es) in 43 pairs of colorectal cancer (CRC) samples. M3Es are widely distributed in CRC and averagely possess around 10% of total active enhancers. We identify 1322 gain variant m3Es and 367 lost variant m3Es in CRC. The target genes of the gain m3Es are enriched in immune response pathways. We experimentally prove that repression of CBX8 and RPS6KA5 m3Es inhibits target gene expression in CRC. Furthermore, we find histone methyltransferase MLL1 is responsible for depositing H3K4me3 on the identified Vm3Es. We demonstrate that the transcription factor AP1/JUN interacts with MLL1 and regulates m3E activity. Application of a small chemical inhibitor for MLL1 activity, OICR-9429, represses target gene expression of the identified Vm3Es, enhances anti-tumor immunity and inhibits CRC growth in an animal model. CONCLUSIONS: Taken together, our study illustrates the genome-wide landscape and the regulatory mechanisms of m3Es in CRC, and reveals potential novel strategies for cancer treatment.

Role of intramembrane charged residues in the quality control of unassembled T-cell receptor alpha-chains at the endoplasmic reticulum.

Endoplasmic reticulum (ER)-associated degradation (ERAD) of unassembled T-cell receptor alpha-chain (TCRalpha) is reliant on the presence of two basic residues in the transmembrane (TM) segment of TCRalpha. The precise role of these residues in ER quality control is unclear. Here, we show that a TCRalpha mutant lacking these intramembrane charged residues has a tendency to form homooligomers through an interchain disulfide bond that involves a specific pair of cysteine residues. Covalent oligomerization of TCRalpha appears to stabilize it at the ER membrane. The presence of a single lysine residue at specific positions within the TCRalpha TM domain abolishes its oligomerization and causes its rapid degradation. Conversely, when TCRalpha oligomerization is induced by a bivalent compound, the degradation of TCRalpha is inhibited. Together, these results suggest that the intramembrane charged residues in TCRalpha do not function as a signal for substrate recognition in ERAD. Instead, their primary role is to reduce TCRalpha oligomerization, maintaining it in a retrotranslocation-competent state. Our results also suggest that the ERAD machinery is inefficient when coping with oligomerized substrates, indicating a requirement for chaperone-mediated protein disassembly in the ER lumen prior to retrotranslocation.

Mechanistic insights into active site-associated polyubiquitination by the ubiquitin-conjugating enzyme Ube2g2.

Lys-48-linked polyubiquitination regulates a variety of cellular processes by targeting ubiquitinated proteins to the proteasome for degradation. Although polyubiquitination had been presumed to occur by transferring ubiquitin molecules, one at a time, from an E2 active site to a substrate, we recently showed that the endoplasmic reticulum-associated RING finger ubiquitin ligase gp78 can mediate the preassembly of Lys-48-linked polyubiquitin chains on the catalytic cysteine of its cognate E2 Ube2g2 and subsequent transfer to a substrate. Active site-linked polyubiquitin chains are detected in cells on Ube2g2 and its yeast homolog Ubc7p, but how these chains are assembled is unclear. Here, we show that gp78 forms an oligomer via 2 oligomerization sites, one of which is a hydrophobic segment located in the gp78 cytosolic domain. We further demonstrate that a gp78 oligomer can simultaneously associate with multiple Ube2g2 molecules. This interaction is mediated by a novel Ube2g2 surface distinct from the predicted RING binding site. Our data suggest that a large gp78-Ube2g2 heterooligomer brings multiple Ube2g2 molecules into close proximity, allowing ubiquitin moieties to be transferred between neighboring Ube2g2s to form active site-linked polyubiquitin chains.

Multifaceted regulation of enhancers in cancer.

Enhancer is one kind of cis-elements regulating gene transcription, whose activity is tightly controlled by epigenetic enzymes and histone modifications. Active enhancers are classified into typical enhancers, super-enhancers and over-active enhancers, according to the enrichment and location of histone modifications. Epigenetic factors control the level of histone modifications on enhancers to determine their activity, such as histone methyltransferases and acetylases. Transcription factors, cofactors and mediators co-operate together and are required for enhancer functions. In turn, abnormalities in these trans-acting factors affect enhancer activity. Recent studies have revealed enhancer dysregulation as one of the important features for cancer. Variations in enhancer regions and mutations of enhancer regulatory genes are frequently observed in cancer cells, and altering the activity of onco-enhancers is able to repress oncogene expression, and suppress tumorigenesis and metastasis. Here we summarize the recent discoveries about enhancer regulation in cancer and discuss their potential application in diagnosis and treatment.

Inhibition of histone methyltransferase SETD8 represses DNA virus replication.

Multiple diseases, such as cancer and neural degeneration diseases, are related with the latent infection of DNA viruses. However, it is still difficult to clean up the latent DNA viruses and new anti-viral strategies are critical for disease treatment. Here, we screen a pool of small chemical molecules and identify UNC0379, an inhibitor for histone H4K20 methyltransferase SETD8, as an effective inhibitor for multiple DNA viruses. UNC0379 not only enhances the expression of anti-viral genes in THP-1 cells, but also repress DNA virus replication in multiple cell lines with defects in cGAS pathway. We prove that SETD8 promotes DNA virus replication in a manner dependent on its enzyme activity. Our results further indicated that SETD8 is required for PCNA stability, one factor critical for viral DNA replication. Viral infection stimulates the interaction between SETD8 and PCNA and thus enhances PCNA stability and viral DNA replication. Taken together, our study reveals a new mechanism for regulating viral DNA replication and provides a potential strategy for treatment of diseases related with DNA viruses.