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Sequences of primers used for RT-PCR. Sequences of siRNA.

Figure S1. sCLU contributes to DEC1 inhibition on DNA damage induced cell death. A, Western blot samples were prepared from MCF7-DEC1-6 cells transfected with scramble siRNA or sCLU siRNA for 48 hours then uninduced or induced to express DEC1 for 12 hours along with mock or CPT (1 mM) treatment for 48 hours. B, DNA histogram analysis was performed with MCF7-DEC1-6 cells. The assay is described in Experimental Procedures (mean SD; n=3). Data are representative of three independent experiments performed in triplicate.

Supplementary Methods, Figures 1-4 from The G Protein–Coupled Receptor 87 Is Necessary for p53-Dependent Cell Survival in Response to Genotoxic Stress

Influenza A viruses (IAVs) rely on viral ribonucleoprotein (vRNP) complexes to control transcription and replication. Each vRNP consists of one viral genomic RNA segment associated with multiple nucleoproteins (NP) and a trimeric IAV RNA polymerase complex. Previous studies showed that post-translational modifications of vRNP components, such as NP, by host factors would in turn affect the IAV life cycle or modulate host anti-viral response. In this study, we found host E3 ubiquitin ligase Pirh2 interacted with NP and mediated short-chain ubiquitination of NP at lysine 351, which suppressed NP-PB2 interaction and vRNP formation. In addition, we showed that knockdown of Pirh2 promoted IAV replication, whereas overexpression of Pirh2 inhibited IAV replication. However, Pirh2-ΔRING lacking E3 ligase activity failed to inhibit IAV infection. Moreover, we showed that Pirh2 had no effect on the replication of a rescued virus, WSN-K351R, carrying lysine-to-arginine substitution at residue 351. Interestingly, by analyzing human and avian IAVs from 2011 to 2020 in influenza research databases, we found that 99.18% of 26 977 human IAVs encode lysine, but 95.3% of 9956 avian IAVs encode arginine at residue 351 of NP protein. Consistently, knockdown of Pirh2 failed to promote propagation of two avian-like influenza viruses, H9N2-W1 and H9N2-C1, which naturally encode arginine at residue 351 of NP. Taken together, we demonstrated that Pirh2 is a host factor restricting IAV infection by modulating short-chain ubiquitination of NP. Meanwhile, it is noteworthy that residue 351 of NP targeted by Pirh2 may associate with the evasion of human anti-viral response against avian-like influenza viruses.

Bovine herpesvirus type 1 (BHV-1) is a virus of causing bovine respiratory disease that poses significant threat to the cattle industry. The prevalence of BHV-1 has recently increased in China. However, information about the prevalent isolates is scarce. In this study, we identified a novel strain of BHV-1, referred to as BHV SHJS, isolated from nasal swabs of Holstein cows in 2020 of China. The genome of BHV SHJS is 1,35,102 bp in length and highly similar to SP1777 (KM258883.1) strain with an identity of 99.64 % . Mutations, insertions, or deletions mainly occur in UL12, UL19, UL27, UL37, UL42, UL44, UL46, UL47, US6, US7, and US8 relative to the different genomic coordinates. Phylogenetic analyses of immunogenic gene (gB, gC, and gD) revealed that BHV SHJS and other China outbreak strains were displayed on different branches with commonly used vaccine strains. Recombination analysis shown that BHV SHJS could persist even with BoviShield IBR MLV and Arsenal IBR MLV vaccines, suggesting that these vaccines should not completely prevent BHV SHJS infection. These analyses partially explain why the vaccines only partially protect against BHV-1 outbreaks in China. In conclusion, we identified a novel and potential recombinant BHV-1 strain and for the first time characterized BHV-1 genome with a different evolutionary origin from that of known strains prevalent in China. This study will enrich our knowledge regarding BHV outbreak strains in China and contribute to the prevention and pathogenic studies of BHV-1.

African swine fever (ASF) is a lethal hemorrhagic disease that affects domestic pigs and wild boars. There is no medication available for ASF to date. The ability to mount antigen-specific responses to viral vectored CP312R makes it a crucial potential target for designing vaccines or drugs. This study determined the crystal structure of ASFV CP312R at 2.32 Å and found it to be a monomer with a single-stranded DNA binding core domain with a clear five-strands β-barrel OB-fold architecture. Electrophoretic mobility shift assay and size-exclusion chromatography characterization assay further confirmed the single-stranded DNA (ssDNA)-binding property of ASFV CP312R. This study revealed the structure and preliminary ssDNA interaction mechanisms of ASFV CP312R, providing new clues for developing new antiviral strategies.

African Swine Fever (ASF) is a highly contagious viral haemorrhagic disease of swine, leading to enormous economic losses in the swine industry. However, vaccines and drugs to treat ASF have yet to be developed. African swine fever virus (ASFV) encodes more than 150 proteins, but 50% of them have unknown functions. Here, we present the crystal structure of the ASFV I73R protein at a resolution of 2.0 Å. Similar search tools based solely on amino acid sequence shows that it has no relationships to any proteins of known function. Interestingly, the overall structure of the I73R protein shares a winged helix-turn-helix fold, structural similarity with the Z-DNA binding domain (Zα). In accordance with this result, the I73R is capable of binding to a CpG repeats DNA duplex, which has a high propensity for forming Z-DNA during the DNA binding assays. In addition, the I73R protein was shown to be expressed at both early and late stages of ASFV post-infection in PAM cells as an 8.9 kDa protein. Immunofluorescence studies revealed that the I73R protein is expressed in the nucleus at early times post-infection and gradually translocated from the nucleus to the cytoplasm. Taken together, these data indicate that the I73R could be a member of Zα family that is important in host-pathogen interaction, which paves the way for the design of inhibitors to target this severe pathogen. Further exploring the biological role of I73R during ASFV infection in vitro and in vivo will provide new clues for development of new antiviral strategies.

Pseudorabies virus (PRV), the causative agent of Aujeszky's disease, has resulted in substantial economic losses in the swine industry worldwide. Previous reports have shown that the PRV variant is responsible for the Pseudorabies outbreaks in Bartha-K61-vaccinated farms in China. However, there is limited information about the evolution of recombination of the PRV variant. Here, we isolated two PRV variants from a Bartha-K61-vaccinated swine farm, named them the JSY7 and JYS13 strains, analysed their complete genomic sequences and evaluated pathogenicity. As results, the JSY7 and JSY13 strains showed different cytopathic effects and plaque sizes. The JSY7 and JSY13 strains had the same Aspartate insertions in the gE protein as other PRV variants. The JSY7 and JSY13 strains were clustered into the same clade based on a genomic phylogenetic analysis. However, the JSY7 strain was relatively close to recent PRV isolates in China, while the JSY13 strain was more closely related to earlier PRV isolates. Interestingly, the gC gene phylogenetic tree showed that the JSY7 strain belonged to genotype II lineage 3, while the JSY13 strain belonged to genotype I and is the same branch with the Bartha strain. Furthermore, the PRV variants were relatively distant from the Bartha strain in the phylogenetic analysis of the gB, gC and gD genes. Importantly, a recombination analysis showed that the JSY13 strain might be a natural recombinant between the minor parental genotype I Bartha strain and the major parental genotype II JSY7 strain. Finally, we also found that the JSY13 strain showed a moderate virulence compared to the JSY7 strain in mice. Taken together, our data provide direct evidence for genomic recombination of PRV in nature, which may play an important role in the evolution and virulence of PRV. This discovery suggests that live PRV vaccine can act as genetic donors for genomic recombination.

The DNA damage response (DDR) pathway is critical for maintaining genomic integrity and sustaining organismal development. Viruses can either utilize or circumvent the DDR to facilitate their replication. Pseudorabies virus (PRV) infection was shown to induce apoptosis via stimulating DDR. However, the underlying mechanisms have not been fully explored to date. This study showed that PRV infection robustly activates the ATM and DNA-PK signaling pathways shortly after infection. However, inhibition of ATM, but not DNA-PK, could dampen PRV replication in cells. Importantly, we found that PRV-encoded serine/threonine kinase UL13 interacts with and subsequently phosphorylates H2AX. Furthermore, we found that UL13 deletion largely attenuates PRV neuroinvasiveness and virulence in vivo. In addtion, we showed that UL13 contributes to H2AX phosphorylation upon PRV infection both in vitro and in vivo, but does not affect ATM phosphorylation. Finally, we showed that knockdown of H2AX reduces PRV replication, while this reduction can be further enhanced by deletion of UL13. Taken together, we conclude that PRV-encoded kinase UL13 regulates DNA damage marker γH2AX and UL13-mediated H2AX phosphorylation plays a pivotal role in efficient PRV replication and progeny production.

The NF-κB pathway is an essential signalling cascade in the defence against viral infections, including African swine fever virus (ASFV) infection. ASFV encodes more than 151 proteins via its own transcription machinery and possesses a great capacity to evade or subvert antiviral innate immune responses. Although some of these viral proteins have been reported, many remain unknown. Here, we show that pD345L, an ASFV-encoded lambda-like exonuclease, acts as an inhibitor of cGAS/STING-mediated NF-κB signalling by blocking the IkappaB kinase (IKKα/β) activity. Specifically, we showed that overexpression of pD345L suppresses cGAS/STING-induced IFNβ and NF-κB activation, resulting in decreased transcription of IFNβ and several proinflammatory cytokines, including IL-1α, IL-6, IL-8, and TNFα. In addition, we showed that pD345L acts at or downstream of IKK and upstream of p65. Importantly, we found that pD345L associates with the KD and HLH domains of IKKα and the LZ domain of IKKβ and thus interrupts their kinase activity towards the downstream substrate IκBα. Finally, we showed that pD345L-mediated inhibition of NF-κB signalling was independent of its exonuclease activity. Considering these results collectively, we concluded that pD345L blocks IKKα/β kinase activity via protein–protein interactions and thus disrupts cGAS/STING-mediated NF-κB signalling.

NF-κB is a critical transcription factor in immediate early viral infection, including African swine fever virus (ASFV), playing an important role in inflammation response and expression of antiviral genes. ASFV encodes for more than 151 proteins by its own transcription machinery and possesses a great capacity to evade or subvert antiviral innate immune responses. A couple of such viral proteins have been reported, but many remain unknown. Here, we showed that pD345L, an ASFV-encoded lambda-like exonuclease, is an inhibitor of cGAS/STING mediated NF-κB signaling by blocking IKKα/β kinase activity. Specifically, we showed that overexpression of pD345L suppresses cGAS/STING induced IFNβ and NF-κB activation, resulting in decreased transcription of IFNβ and several pro-inflammatory cytokines, including IL-1α, IL-6, IL-8, and TNFα. In addition, we showed that pD345L targeted at or downstream of IKK and upstream of p65. Importantly, we found that pD345L associates with KD and HLH domains of IKKα and LZ domain of IKKβ, and thus interrupts their kinase activity on downstream substrate IκBα. Finally, we showed that pD345L inhibition of NF-κB signaling was independent of its exonuclease activity. Taken together, we concluded that pD345L blocks IKK α/β kinase activity by protein-protein interaction and thus disrupts cGAS/STING mediated NF-κB signaling.ImportanceAfrican Swine Fever (ASF) is one of the most devastating and economically significant swine diseases caused by African swine fever virus (ASFV). Since expanding of ASFV affected areas into Asian countries, especially China, an effective vaccine is urgently needed more than ever. Therefore, it is of great significance to understand the interaction between ASFV infection and host immune responses. The NF-κB signaling plays a central role in innate and acquired immune responses. Activation of IκB kinase (IKK) complex is a key step of both canonical and non-canonical NF-κB pathways, and is commonly targeted by different viruses. But no ASFV protein has been shown to regulate IKK yet. In this study, we demonstrated that pD345L blocks IKKα/β kinase activity by protein-protein interaction and thus disrupts cGAS/STING mediated NF-κB signaling. It has been shown that conventional vaccine development approaches are proven to be inapplicable to ASFV. Neither subunit nor DNA vaccine provides efficient protection. Gene deleted live-attenuated vaccine candidates render adequate protection, but establishment of chronic or persistent infection in vaccinated animals and risk of recombination with filed strains are key challenges. To overcome these, one potential strategy would be generation of replication-defective viruses. As a lambda-like exonuclease, pD345L plays a critical role in ASFV replication and ASFV deficient in D345L cannot be rescued. Given the dual role of pD345L in virus replication and immune evasion, it may serve as a potential target for replication-defective virus vaccine development.

Graphene oxide (GO) is recognized as a promising antibacterial material that is expected to be used to prepare a new generation of high-efficiency antibacterial coatings. The propensity of GO to agglomeration makes it difficult to apply it effectively. A new method of preparing GO-loaded nickel (GNC) with excellent antibacterial property is proposed in this paper. In this work, GNC was prepared on a titanium sheet by magnetic field-assisted scanning jet electrodeposition. The massive introduction of GO on the coating was proven by energy disperse spectroscopy and Raman spectroscopy. The antibacterial performance of GNC was proven by agar plate assessment and cell living/dead staining. The detection of intracellular reactive oxygen species (ROS) and the concentration of nickel ions, indicate that the antibacterial property of GNC are not entirely derived from the nickel ions released by the coating and the intracellular ROS induced by nickel ions, but rather are due to the synergistic effect of nickel ions and GO.

Porcine epidemic diarrhea virus (PEDV) is an α-coronavirus causing severe diarrhea and high mortality rates in suckling piglets and posing significant economic impact. PEDV replication is completed and results in a large amount of RNA in the cytoplasm. Stress granules (SGs) are dynamic cytosolic RNA granules formed under various stress conditions, including viral infections. Several previous studies suggested that SGs were involved in the antiviral activity of host cells to limit viral propagation. However, the underlying mechanisms are poorly understood. This study aimed to delineate the molecular mechanisms regulating the SG response to PEDV infection. SG formation is induced early during PEDV infection, but as infection proceeds, this ability is lost and SGs disappear at late stages of infection (>18 h postinfection). PEDV infection resulted in the cleavage of Ras-GTPase-activating protein-binding protein 1 (G3BP1) mediated by caspase-8. Using mutational analysis, the PEDV-induced cleavage site within G3BP1 was identified, which differed from the 3C protease cleavage site previously identified. Furthermore, G3BP1 cleavage by caspase-8 at D168 and D169 was confirmed in vitro as well as in vivo. The overexpression of cleavage-resistant G3BP1 conferred persistent SG formation and suppression of viral replication. Additionally, the knockdown of endogenous G3BP1 abolished SG formation and potentiated viral replication. Taken together, these data provide new insights into novel strategies in which PEDV limits the host stress response and antiviral responses and indicate that caspase-8-mediated G3BP1 cleavage is important in the failure of host defense against PEDV infection. IMPORTANCE Coronaviruses (CoVs) are drawing extensive attention again since the outbreaks of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in 2019. CoVs are prone to variation and own the transmission capability by crossing the species barrier resulting in reemergence. How CoVs manipulate the antiviral responses of their hosts needs to be explored. Overall, the study provides new insight into how porcine epidemic diarrhea virus (PEDV) impaired SG assembly by targeting G3BP1 via the host proteinase caspase-8. These findings enhanced the understanding of PEDV infection and might help identify new antiviral targets that could inhibit viral replication and limit the pathogenesis of PEDV.

The neddylation pathway belongs post-translational modifications and plays important roles in regulating viral infection and replication. To address the relationship of influenza A virus with the neddylation modification pathway, we demonstrate that IAV infection in A549 cells can activate the neddylation modification pathway to increase virus growth and enhance the expression of pro-inflammatory cytokines to increase pathogenicity. The pre-treatment of Nedd8-activating enzyme subunit 1 (NAE1)-specific inhibitor, MLN4924, interferes with Nedd8 conjugation and NF-κB activity. MLN4924 exhibited pronounced antiviral activity against different subtypes of influenza A virus, including classical H1N1 (PR8), H9N2 subtype, and pandemic H1N1 2009 (pdmH1N1) viruses. Through the inhibition of the CRL/NF-κB pathway, MLN4924 could significantly suppress the expression levels of pro-inflammatory cytokines induced by IAVs. These findings suggest that MLN4924 can be developed as a novel antiviral therapy for influenza infection for anti-viral efficacy and anti-inflammation activity.

This work studies the antibacterial and anti-corrosion properties of nickel-graphene oxide (Ni-GO) composite coating and nickel-reduced graphene oxide (Ni-rGO) composite coating deposited by magnetic field assisted scanning jet electrodeposition on manganese steel. Agar plate assessment and cell live/dead staining were used to evaluate the antibacterial property of the two types of composite coatings. The Ni-GO and Ni-rGO composite coatings can effectively improve the antibacterial property of the substrate. The Ni-GO composite coating deposited with graphene oxide (GO) concentration of 0.5 g/L showed the most excellent antibacterial ability, and its antibacterial rate against E. coli and S. aureus within 2 h were 100% and 93%, respectively. Potentiodynamic polarization measurements indicated that the protective efficiency of the two types of composite coatings on the manganese steel reaches 99% after being immersed in 3.5 wt% NaCl solution for 7 days. The combined effect of barrier protection from GO-based materials (GO, rGO) and corrosion products effectively inhibits the corrosion behavior of the two types of composite coatings.

Health differences between breast- and formula-fed infants have long been apparent despite great efforts in improving the function of baby formula by adjusting the levels of various milk nutritional components. However, the N-glycome, a type of oligosaccharide decorating a diverse range of proteins, has not been extensively studied in milk regarding its biological function. In this study, the anti-pathogenic function of the enzymatically released human and bovine milk N-glycome against 5 food-borne pathogens was investigated. The human milk N-glycome showed significantly higher activity than bovine milk. After enzymatic defucosylation of human and bovine N-glycan pool, UHPLC peak shifts were observed in both suggesting heavy fucosylation of samples. Furthermore, the anti-pathogenic activity of the defulosylated N-glycome decreased significantly, and the significance of functional difference between the two almost disappeared. This result indicates the essential role of fucosylation for the anti-pathogenic function of the milk N-glycome, especially in human milk.

Novel sialic acid scaffolds have great significance in the development of influenza neuraminidase inhibitors. Here the enzymatic synthesis of a wide range of 2-keto-3-deoxy-D-glycero-D-galacto-nononic acid (KDN) analogues via aldol addition of pyruvate to D-mannose, D-glucose, D-galactose, 2-deoxy-D-glucose, D-arabinose, L-arabinose and L-rhamnose using a previously unstudied N-acetylneuraminic acid (Neu5Ac) aldolase derived from the bacterium Dyadobacter fermentas was exemplified. Several of the synthesized KDN analogues showed comparable or better inhibitory activity than unstudied Neu5Ac against the mutated influenza neuraminidases (A/California/04/2009 and A/Anhui/1/2005), which both show resistance to Neu5Ac-based neuraminidase inhibitors, demonstrating that these compounds are promising templates for the development of anti-influenza drugs.

Porcine Reproductive and Respiratory Syndrome (PRRS), which is caused by Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) infection, has caused substantial economic losses for the global swine industry. To date, there are limited commercially available measures to control the spread of PRRSV. The available vaccines are unstable and there is no anti-PRRSV therapeutic available. Therefore, this study designed a novel recombinant antiviral protein that included a novel polypeptide that binds to the PRRSV polymerase (p9), the transduction ability of the HIV TAT, and the nucleotide degradation activity of interferon stimulated gene 20 (ISG20). The recombinant proteins TAT-p9-ISG20 and p9-ISG20 were expressed in MARC-145 cells by transient transfection and then tested for antiviral activity and entry efficiency. The p9-ISG20 construct had greater antiviral activity than either p9 alone (1.37-fold) or ISG20 alone (1.9-fold). Addition of the HIV TAT protein increased the entry efficiency of p9-ISG20 by 1.57-fold, which was associated with increased anti-viral activity (1.52-fold) compared to P9-ISG20. In summary, TAT-p9-ISG20 achieved a synergistic effect by combining three different antiviral proteins and may be a novel PRRSV therapeutic platform.

Oncogenic virus replication often leads to genomic instability, causing DNA damage and inducing the DNA damage response (DDR) pathway. The DDR pathway is a cellular pathway that senses DNA damage and regulates the cell cycle to maintain genomic stability. Therefore, the DDR pathway is critical for the viral lifecycle and tumorigenesis. Marek’s disease virus (MDV), an alphaherpesvirus that causes lymphoma in chickens, has been shown to induce DNA damage in infected cells. However, the interaction between MDV and the host DDR is unclear. In this study, we observed that MDV infection causes DNA strand breakage in chicken fibroblast (CEF) cells along with an increase in the DNA damage markers p53 and p21. Interestingly, we showed that phosphorylation of STAT3 was increased during MDV infection, concomitantly with a decrease of Chk1 phosphorylation. In addition, we found that MDV infection was enhanced by VE-821, an ATR-specific inhibitor, but attenuated by hydroxyurea, an ATR activator. Moreover, inhibition of STAT3 phosphorylation by Stattic eliminates the ability of MDV to inhibit Chk1 phosphorylation. Finally, we showed that MDV replication was decreased by Stattic treatment. Taken together, these results suggest that MDV disables the ATR-Chk1 pathway through STAT3 activation to benefit its replication. IMPORTANCE MDV is used as a biomedical model to study virus-induced lymphoma due to the similar genomic structures and physiological characteristics of MDV and human herpesviruses. Upon infection, MDV induces DNA damage, which may activate the DDR pathway. The DDR pathway has a dual impact on viruses because it manipulates repair and recombination factors to facilitate viral replication and also initiates antiviral action by regulating other signaling pathways. Many DNA viruses evolve to manipulate the DDR pathway to promote virus replication. In this study, we identified a mechanism used by MDV to inhibit ATR-Chk1 pathways. ATR is a cellular kinase that responds to broken single-stranded DNA, which has been less studied in MDV infection. Our results suggest that MDV infection activates STAT3 to disable the ATR-Chk1 pathway, which is conducive to viral replication. This finding provides new insight into the role of STAT3 in interrupting the ATR-Chk1 pathway during MDV replication.

Influenza A virus (IAV) continues to threaten global public health due to drug resistance and the emergence of frequently mutated strains. Thus, it is critical to find new strategies to control IAV infection. Here, we discover one host protein, HDAC6, that can inhibit viral RNA polymerase activity by deacetylating PA and thus suppresses virus RNA replication and transcription. Previously, it was reported that IAV can utilize the HDAC6-dependent aggresome formation mechanism to promote virus uncoating, but HDAC6-mediated deacetylation of α-tubulin inhibits viral protein trafficking at late stages of the virus life cycle. These findings together will contribute to a better understanding of the role of HDAC6 in regulating IAV infection. Understanding the molecular mechanisms of HDAC6 at various periods of viral infection may illuminate novel strategies for developing antiviral drugs., The life cycle of influenza A virus (IAV) is modulated by various cellular host factors. Although previous studies indicated that IAV infection is controlled by HDAC6, the deacetylase involved in the regulation of PA remained unknown. Here, we demonstrate that HDAC6 acts as a negative regulator of IAV infection by destabilizing PA. HDAC6 binds to and deacetylates PA, thereby promoting the proteasomal degradation of PA. Based on mass spectrometric analysis, Lys(664) of PA can be deacetylated by HDAC6, and the residue is crucial for PA protein stability. The deacetylase activity of HDAC6 is required for anti-IAV activity, because IAV infection was enhanced due to elevated IAV RNA polymerase activity upon HDAC6 depletion and an HDAC6 deacetylase dead mutant (HDAC6-DM; H216A, H611A). Finally, we also demonstrate that overexpression of HDAC6 suppresses IAV RNA polymerase activity, but HDAC6-DM does not. Taken together, our findings provide initial evidence that HDAC6 plays a negative role in IAV RNA polymerase activity by deacetylating PA and thus restricts IAV RNA transcription and replication. IMPORTANCE Influenza A virus (IAV) continues to threaten global public health due to drug resistance and the emergence of frequently mutated strains. Thus, it is critical to find new strategies to control IAV infection. Here, we discover one host protein, HDAC6, that can inhibit viral RNA polymerase activity by deacetylating PA and thus suppresses virus RNA replication and transcription. Previously, it was reported that IAV can utilize the HDAC6-dependent aggresome formation mechanism to promote virus uncoating, but HDAC6-mediated deacetylation of α-tubulin inhibits viral protein trafficking at late stages of the virus life cycle. These findings together will contribute to a better understanding of the role of HDAC6 in regulating IAV infection. Understanding the molecular mechanisms of HDAC6 at various periods of viral infection may illuminate novel strategies for developing antiviral drugs.

Background Marek’s disease virus (MDV) resides in the genus Mardivirus in the family Herpesviridae. MDV is a highly contagious virus that can cause neurological lesions, lymphocytic proliferation, immune suppression, and death in avian species, including Galliformes (chickens, quails, partridges, and pheasants), Strigiformes (owls), Anseriformes (ducks, geese, and swans), and Falconiformes (kestrels). Case presentation In 2015, two red-crowned cranes died in Nanjing (Jiangsu, China). It was determined that the birds were infected with Marek’s disease virus by histopathological examination, polymerase chain reaction (PCR), gene sequencing and sequence analysis of tissue samples from two cranes. Gross lesions included diffuse nodules in the skin, muscle, liver, spleen, kidney, gizzard and heart, along with liver enlargement and gizzard mucosa hemorrhage. Histopathological assay showed that infiltrative lymphocytes and mitotic figures existed in liver and heart. The presence of MDV was confirmed by PCR. The sequence analysis of the Meq gene showed 100% identity with Md5, while the VP22 gene showed the highest homology with CVI988. Furthermore, the phylogenetic analysis of the VP22 and Meq genes suggested that the MDV (from cranes) belongs to MDV serotype 1. Conclusion We describe the first molecular detection of Marek’s disease in red-crowned cranes based on the findings previously described. To our knowledge, this is also the first molecular identification of Marek’s disease virus in the order Gruiformes and represents detection of a novel MDV strain.

Differentiated embryonic chondrocyte expressed gene 1 (DEC1, also known as Sharp2/Stra13/BHLHE40) is a basic helix–loop–helix transcription factor that plays an important role in circadian rhythms, cell proliferation, apoptosis, cellular senescence, hypoxia response, and epithelial-to-mesenchymal transition of tumor cells. Secretory clusterin (sCLU) is a cytoprotective protein that guards against genotoxic stresses. Here, clusterin (CLU) was identified as a novel target gene of DEC1 and suppresses DNA damage–induced cell death in tumor cells. Mechanistically, based on chromatin immunoprecipitation and luciferase assays, DEC1 binds to and activates the promoter of the CLU gene. DEC1 and DNA-damaging agents induce sCLU expression, whereas DEC1 knockdown decreases the expression of sCLU upon DNA damage. Moreover, the data demonstrate that DEC1 inhibits, whereas sCLU knockdown enhances, DNA damage–induced cell death in MCF7 breast cancer cells. Given that DEC1 and sCLU are frequently overexpressed in breast cancers, these data provide mechanistic insight into DEC1 as a prosurvival factor by upregulating sCLU to reduce the DNA damage–induced apoptotic response. Together, this study reveals sCLU as a novel target of DEC1 which modulates the sensitivity of the DNA damage response. Implications: DEC1 and sCLU are frequently overexpressed in breast cancer, and targeting the sCLU-mediated cytoprotective signaling pathway may be a novel therapeutic approach. Mol Cancer Res; 16(11); 1641–51. ©2018 AACR.

Japanese encephalitis virus (JEV) is an arthropod-borne flavivirus prevalent in Asia and the Western Pacific and is the leading cause of viral encephalitis. JEV is maintained in a transmission cycle between mosquitoes and vertebrate hosts, but the molecular mechanisms by which the mosquito vector participates in transmission are unclear. We investigated the expression of all C-type lectins during JEV infection in Aedes aegypti . The C-type lectin mosquito galactose-specific C-type lectin 7 (mosGCTL-7) (VectorBase accession no. AAEL002524) was significantly upregulated by JEV infection and facilitated infection in vivo and in vitro . mosGCTL-7 bound to the N-glycan at N154 on the JEV envelope protein. This recognition of viral N-glycan by mosGCTL-7 is required for JEV infection, and we found that this interaction was Ca 2+ dependent. After mosGCTL-7 bound to the glycan, mosPTP-1 bound to mosGCTL-7, promoting JEV entry. The viral burden in vivo and in vitro was significantly decreased by mosPTP-1 double-stranded RNA (dsRNA) treatment, and infection was abolished by anti-mosGCTL-7 antibodies. Our results indicate that the mosGCTL-7/mosPTP-1 pathway plays a key role in JEV infection in mosquitoes. An improved understanding of the mechanisms underlying flavivirus infection in mosquitoes will provide further opportunities for developing new strategies to control viral dissemination in nature. IMPORTANCE Japanese encephalitis virus is a mosquito-borne flavivirus and is the primary cause of viral encephalitis in the Asia-Pacific region. Twenty-four countries in the WHO Southeast Asia and Western Pacific regions have endemic JEV transmission, which exposes >3 billion people to the risks of infection, although JEV primarily affects children. C-type lectins are host factors that play a role in flavivirus infection in humans, swine, and other mammals. In this study, we investigated C-type lectin functions in JEV-infected Aedes aegypti and Culex pipiens pallens mosquitoes and cultured cells. JEV infection changed the expression of almost all C-type lectins in vivo and in vitro , and mosGCTL-7 bound to the JEV envelope protein via an N-glycan at N154. Cell surface mosPTP-1 interacted with the mosGCTL-7–JEV complex to facilitate virus infection in vivo and in vitro . Our findings provide further opportunities for developing new strategies to control arbovirus dissemination in nature.

Ferredoxin reductase (FDXR), a target of p53, modulates p53-dependent apoptosis and is necessary for steroidogenesis and biogenesis of iron-sulfur clusters. To determine the biological function of FDXR, we generated a Fdxr-deficient mouse model and found that loss of Fdxr led to embryonic lethality potentially due to iron overload in developing embryos. Interestingly, mice heterozygous in Fdxr had a short life span and were prone to spontaneous tumors and liver abnormalities, including steatosis, hepatitis, and hepatocellular carcinoma. We also found that FDXR was necessary for mitochondrial iron homeostasis and proper expression of several master regulators of iron metabolism, including iron regulatory protein 2 (IRP2). Surprisingly, we found that p53 mRNA translation was suppressed by FDXR deficiency via IRP2. Moreover, we found that the signal from FDXR to iron homeostasis and the p53 pathway was transduced by ferredoxin 2, a substrate of FDXR. Finally, we found that p53 played a role in iron homeostasis and was required for FDXR-mediated iron metabolism. Together, we conclude that FDXR and p53 are mutually regulated and that the FDXR-p53 loop is critical for tumor suppression via iron homeostasis.

The cellular DNA sensor cGMP-AMP synthase (cGAS) detects cytosolic viral DNA via the stimulator of interferon genes (STING) to initiate innate antiviral response. Herpesviruses are known to target key immune signaling pathways to persist in an immune-competent host. Marek’s disease virus (MDV), a highly pathogenic and oncogenic herpesvirus of chickens, can antagonize host innate immune responses to achieve persistent infection. With a functional screen, we identified five MDV proteins that blocked beta interferon (IFN-β) induction downstream of the cGAS-STING pathway. Specifically, the MDV major oncoprotein Meq impeded the recruitment of TANK-binding kinase 1 and IFN regulatory factor 7 (IRF7) to the STING complex, thereby inhibiting IRF7 activation and IFN-β induction. Meq overexpression markedly reduced antiviral responses stimulated by cytosolic DNA, whereas knockdown of Meq heightened MDV-triggered induction of IFN-β and downstream antiviral genes. Moreover, Meq-deficient MDV induced more IFN-β production than wild-type MDV. Meq-deficient MDV also triggered a more robust CD8+ T cell response than wild-type MDV. As such, the Meq-deficient MDV was highly attenuated in replication and lymphoma induction compared to wild-type MDV. Taken together, these results revealed that MDV evades the cGAS-STING DNA sensing pathway, which underpins the efficient replication and oncogenesis. These findings improve our understanding of the virus-host interaction in MDV-induced lymphoma and may contribute to the development of novel vaccines against MDV infection., Author summary Marek’s disease virus (MDV) is an avian oncogenic herpesvirus that causes a fatal disease in poultry worldwide. Chickens infected with MDV become more susceptible to secondary viral or bacterial infections. However, the mechanisms of MDV-induced immunosuppression and tumorigenesis remain largely unknown. The cGAS-STING pathway is crucial for innate immune responses against both microbial pathogens and intrinsic tumors. Here we identified the MDV oncoprotein, Meq, as an inhibitor of the cGAS-STING DNA-sensing pathway. Mechanistically, Meq interacted with STING and IRF7, and impaired the recruitment of TBK1 and IRF7 to the STING complex, thus inhibiting IRF7 activation and IFN-β induction. Loss of Meq potently enhanced innate immune response, while impaired the replication and oncogenesis of MDV in chickens. Our findings reveal an important mechanism of immune evasion of MDV, instructing us on the virus-host interaction in MDV-induced lymphoma and potential new means to develop MDV vaccine.

Tumor suppressor p53 maintains genome stability by regulating diverse cellular functions including cell cycle arrest, apoptosis, senescence and metabolic homeostasis. Mutations in the p53 gene occur in almost all human cancers with a frequency of up to 80%. However, it is only 20% in breast cancers, 18% in endometrial cancers and 1.5% in cervical cancers. Estrogen receptor alpha (ERα) plays a pivotal role in hormone-dependent cancer development and the status of ERα is used for designing treatment strategy and for prognosis. A closer look at the cross-talk between p53 and ERα has revealed that their activities are mutually regulated. This review will summarize the current body of knowledge on p53, ERα and ERβ in cancer. Clinical correlations between estrogen receptors and p53 status have also been reported. Thus, this review will discuss the relationship between p53 and ERs at both the molecular and clinical levels.

In response to genotoxic stress, the p53 tumor suppressor induces target genes for cell cycle arrest, apoptosis, and DNA repair. Although p53 is the most commonly mutated gene in all human cancers, it is only mutated in about 20% of breast cancers. 70% of all breast cancer cases are estrogen receptor (ER)-positive and express ERα. ER-positive breast cancer generally indicates good patient prognosis and treatment responsiveness with antiestrogens, such as tamoxifen. However, ER-positive breast cancer patients can experience loss or a reduction in ERα, which is associated with aggressive tumor growth, increased invasiveness, poor prognosis, and loss of p53 function. Consistent with this, we found that p53 is a target gene of ERα. Specifically, we found that knockdown of ERα decreases expression of p53 and its downstream targets, MDM2 and p21. In addition, we found that ERα activates p53 transcription via binding to estrogen response element half-sites within the p53 promoter. Moreover, we found that loss of ERα desensitizes, whereas ectopic expression of ERα sensitizes, breast cancer cells to DNA damage-induced growth suppression in a p53-dependent manner. Altogether, this study provides an insight into a feedback loop between ERα and p53 and a biological role of p53 in the DNA damage response in ER-positive breast cancers.

The ubiquitin-dependent proteasome system plays a critical role in many cellular processes and pathogenesis of various human diseases, including cancer. Although there are a large number of E3 ubiquitin ligases, the majority are RING-finger type E3s. Pirh2, a target of p53 transcription factor, contains a highly conserved C3H2C3 type RING domain. Importantly, Pirh2 was found to regulate a group of key factors dedicated to the DNA damage response, such as p53, p73, PolH, and c-Myc. Interestingly, Pirh2 was upregulated or downregulated in different types of cancers. These suggest that Pirh2 is implicated in either promoting or suppressing tumor progression in a tissue-dependent manner. This review will focus on the major findings in these studies and discuss the potential to explore Pirh2 as a cancer therapeutic target.

The p73 gene, a homologue of the p53 tumor suppressor, is expressed as TA and ΔN isoforms. TAp73 has similar activity as p53 and functions as a tumor suppressor whereas ΔNp73 has both pro- and anti-survival functions. While p73 is rarely mutated in spontaneous tumors, the expression status of p73 is linked to the sensitivity of tumor cells to chemotherapy and prognosis for many types of human cancer. Thus, uncovering its regulators in tumors is of great interest. Here, we found that Pirh2, a RING finger E3 ubiquitin ligase, promotes the proteasome-dependent degradation of p73. Specifically, we showed that knockdown of Pirh2 up-regulates, whereas ectopic expression of Pirh2 down-regulates, expression of endogenous and exogenous p73. In addition, Pirh2 physically associates with and promotes TAp73 polyubiquitination both in vivo and in vitro. Moreover, we found that p73 can be degraded by both 20 S and 26 S proteasomes. Finally, we showed that Pirh2 knockdown leads to growth suppression in a TAp73-dependent manner. Taken together, our findings indicate that Pirh2 promotes the proteasomal turnover of TAp73, and thus targeting Pirh2 to restore TAp73-mediated growth suppression in p53-deficient tumors may be developed as a novel anti-cancer strategy.

The p63 gene, a member of the p53 family, is expressed as TA and ΔN isoforms. ΔNp63 is the predominant isoform expressed in cells of epithelial origin and frequently overexpressed in cancers. However, what regulates p63 expression is uncertain. Here, we showed that ΔNp63 is regulated by the transcription factor DEC1, a p53 family target. We also showed that the ability of DEC1 to regulate ΔNp63 is enhanced by histone deacetylase (HDAC) inhibitors or knockdown of histone deacetylase 2 (HDAC2). Consistent with this, we found that DEC1 and HDAC2 physically interact and knockdown of HDAC2 leads to increased binding of DEC1 to the ΔNp63 promoter. Interestingly, we found that growth suppression induced by HDAC inhibitors is attenuated by ectopic expression of DEC1 in a ΔNp63-dependent manner. In addition, we showed that ectopic expression of DEC1 inhibits, whereas knockdown of DEC1 promotes, keratinocyte differentiation via modulating ΔNp63 expression. Finally, we showed that DEC1 cooperates with HDAC inhibitors to further decrease keratinocyte differentiation. Together, we conclude that ΔNp63 is a novel target of DEC1 and HDAC2 and modulates the efficacy of HDAC inhibitors in growth suppression and keratinocyte differentiation.

Cellular senescence plays an important role in tumor suppression. p53 tumor suppressor has been reported to be crucial in cellular senescence. However, the underlying mechanism is poorly understood. In this regard, a cDNA microarray assay was performed to identify p53 targets involved in senescence. Among the many candidates is DEC1, a basic helix-loop-helix transcription factor that has been recently shown to be up-regulated in K-ras-induced premature senescence. However, it is not clear whether DEC1 is capable of inducing senescence. Here, we found that DEC1 is a novel target gene of the p53 family and mediates p53-dependent premature senescence. Specifically, we showed that DEC1 is induced by the p53 family and DNA damage in a p53-dependent manner. We also found that the p53 family proteins bind to, and activate, the promoter of the DEC1 gene. In addition, we showed that overexpression of DEC1 induces G(1) arrest and promotes senescence. Moreover, we found that targeting endogenous DEC1 attenuates p53-mediated premature senescence in response to DNA damage. Furthermore, overexpression of DEC1 induces cellular senescence in p53-knockdown cells, albeit to a lesser extent. Finally, we showed that DEC1-induced senescence is p21-independent. Taken together, our data provided strong evidence that DEC1 is one of the effectors downstream of p53 to promote premature senescence.

p21, a cyclin-dependent kinase inhibitor, is necessary for proper control of the cell cycle and premature senescence. Thus, p21 expression needs to be tightly controlled. In this study, we found that Rbm24, an RNA-binding protein and a target gene of the p53 protein, can regulate p21 expression via mRNA stability. Specifically, we showed that Rbm24 is induced by DNA damage and Mdm2 inhibitor Nutlin-3. We also found that p53 protein binds to and activates the promoter of the Rbm24 gene. Moreover, we found that overexpression of Rbm24 increases, whereas knockdown of Rbm24 decreases, p21 mRNA and protein expression. In addition, we demonstrated that overexpression of Rbm24 enhances the half-life of p21 transcript. Consistent with this, we provided evidence that Rbm24 binds to the 3′-untranslated region (3′-UTR) of p21 transcript and an AU/U-rich element in the p21 3′-UTR is necessary for Rbm24 to increase p21 expression. Finally, we showed that the RNA recognition motif in Rbm24 is required for binding to p21 transcript and subsequently for inducing p21 expression. Altogether, we uncovered that Rbm24 is a novel player in the p53 pathway, which may be explored to restore proper cell cycle control in p53-deficient tumors via p21.

P73, a member of the p53 family, plays a critical role in neural development and tumorigenesis. Due to the usage of two different promoters, p73 is expressed as two major isoforms, TAp73 and ΔNp73, often with opposing functions. Here, we reported that transcriptional factor DEC1, a target of the p53 family, exerts a distinct control of TAp73 and ΔNp73 expression. In particular, we showed that DEC1 was able to increase TAp73 expression via transcriptional activation of the TAp73 promoter. By contrast, Np73 transcription was inhibited by DEC1 via transcriptional repression of the ΔNp73 promoter. To further explore the underlying mechanism, we showed that DEC1 was unable to increase TAp73 expression in the absence of HDAC8, suggesting that HDAC8 is required for DEC1 to enhance TAp73 expression. Furthermore, we found that DEC1 was able to interact with HDAC8 and recruit HDAC8 to the TAp73, but not the ΔNp73, promoter. Together, our data provide evidence that DEC1 and HDAC8 in differentially regulate TAp73 and ΔNp73 expression, suggesting that this regulation may lay a foundation for a therapeutic strategy to enhance the chemosensitivity of tumor cells.

p53, a guardian of the genome, exerts its tumor suppression activity by regulating a large number of downstream targets involved in cell cycle arrest, DNA repair, apoptosis, and cellular senescence. Although p53-mediated apoptosis is able to kill cancer cells, a role for cellular senescence in p53-dependent tumor suppression is becoming clear. Mouse studies showed that activation of p53-induced premature senescence promotes tumor regression in vivo. However, p53-mediated cellular senescence also leads to aging-related phenotypes, such as tissue atrophy, stem cell depletion, and impaired wound healing. In addition, several p53 isoforms and two p53 homologs, p63 and p73, have been shown to play a role in cellular senescence and/or aging. Importantly, p53, p63, and p73 are necessary for the maintenance of adult stem cells. Therefore, understanding the dual role the p53 protein family in cancer and aging is critical to solve cancer and longevity in the future. In this chapter, we provide an overview on how p53, p63, p73, and their isoforms regulate cellular senescence and aging.

DNA polymerase eta (PolH), the product of the xeroderma pigmentosum variant (XPV) gene and a Y-family DNA polymerase, plays a pivotal role in translesion DNA synthesis. Loss of PolH leads to early onset of malignant skin cancer in XPV patients and increases UV-induced carcinogenesis. Thus, the pathways by which PolH expression and activity are controlled may be explored as a strategy to prevent UV-induced cancer. In this study, we found that Mdm2, a RING finger E3 ligase, promotes PolH degradation. Specifically, we showed that knockdown of Mdm2 increases PolH expression in both p53-proficient and -deficient cells. In addition, we showed that UV-induced PolH degradation is attenuated by Mdm2 knockdown. In contrast, ectopically expression of Mdm2 decreases PolH expression, which can be abrogated by the proteasome inhibitor MG132. Moreover, we showed that Mdm2 physically associates with PolH and promotes PolH polyubiquitination in vivo and in vitro. Finally, we showed that knockdown of Mdm2 increases the formation of PolH replication foci and decreases the sensitivity of cells to UV-induced lesions in a PolH-dependent manner. Taken together, we uncovered that Mdm2 serves as an E3 ligase for PolH polyubiquitination and proteasomal degradation in cells under the basal condition and in response to UV irradiation.

Cellular senescence is a permanent cell cycle arrest and a potent tumor suppression mechanism. The p53 tumor suppressor is a sequence-specific transcription factor and acts as a central hub sensing various stress signals and activating an array of target genes to induce cell cycle arrest, apoptosis, and senescence. Recent reports showed that restoration of p53 induces premature senescence and tumor regression in mice with hepatocarcinomas or sarcomas. Thus, p53-mediated senescence is capable of eliminating cancer cells in vivo. p63 and p73, two homologues of p53, have similar function in cell cycle arrest and apoptosis. However, the role of p63 and p73 in cellular senescence is elusive. In this review, we will discuss how p53 regulates senescence and future studies about p53 family members in senescence.

p53 regulates an array of target genes, which mediates p53 tumor suppression by inducing cell cycle arrest, apoptosis, and cell survival. G protein–coupled receptors belong to a superfamily of cell surface molecules and are known to regulate cell proliferation, migration, and survival. Here, we found that G protein–coupled receptor 87 (GPR87) was up-regulated by p53 and by DNA damage in a p53-dependent manner. We also found that p53 directly regulated GPR87 potentially via a p53-responsive element in the GPR87 gene. To investigate the role of GPR87 in the p53 pathway, we generated multiple RKO and MCF7 cell lines in that GPR87 can be inducibly overexpressed or knocked down by a tetracycline-inducible system. We found that overexpression of GPR87 had little effect on cell growth. However, GPR87 knockdown sensitized cancer cells to DNA damage–induced growth suppression via enhanced p53 stabilization and activation. Importantly, the prosurvival activity of GPR87 can be reversed by knockdown of p53. Together, our results suggested that GPR87 is essential for p53-dependent cell survival in response to DNA damage. Thus, due to its expression on the cell surface and its role in cell survival, GPR87 may be explored as a novel therapeutic target for cancer treatment and prevention. [Cancer Res 2009;69(15):6049–56]

The p53 tumor suppressor is an essential barrier against cancer, which is evidenced by loss of p53 activity in the majority of human cancers and unexceptionally early onset of tumors in p53-knockout mice and human Li-Fraumeni syndrome patients. p53 is a sequence-specific transcription factor that regulates gene expression via binding to the consensus p53-responsive element (p53-RE). p53 acts as a central signaling coordinator that receives upstream signals, including DNA damage, aberrant oncogenic activity, hypoxia and ribosomal stress, and consequently activates its targets leading to downstream cellular outcomes, including cell cycle arrest, DNA repair, apoptosis and cellular senescence. However, it is not yet clear how p53 dictates a cell to survival vs. death in response to a specific stress. It has been shown that the ability of p53 to regulate target genes can be modulated by posttranslational modifications. For example, acetylation of p53 on lysine 120 by Tip60 is crucial for p53-mediated apoptosis but not cell cycle arrest.1 Consistently, p90, a cofactor of Tip60, is required for Tip60-dependent p53 acetylation and, thus, specifically promotes p53-mediated apoptosis.2 Interestingly, evidence showed that p53 activity is also regulated by binding partners that do not affect p53 modifications. For example, ASPP proteins interact with p53 and enhance the DNA binding and transactivation function of p53 on the promoters of pro-apoptotic genes, such as Bax and PIG-3.3 Similarly, p53β preferentially enhances p53 transcriptional activity on the Bax but not p21 promoter.4 In addition, hCAS/CSE1L, a factor identified in the p53 transcription complex, specifically associates with the PIG-3 promoter to induce p53-mediated apoptosis.5 By contrast, BRCA16 and HZF7 direct p53-dependent transactivation of pro-arrest target genes over pro-apoptotic target genes. Recently, we found that DEC1, a target of the p53 family and a basic helix-loop-helix (bHLH) transcription factor, modulates p53-depedent cell survival vs. cell death in response to genotoxic stress.8 In addition, DEC1 is capable of inducing cell cycle arrest in a p53-independent manner.9 As a transcription factor, DEC1 is able to repress gene expression through class B E-box elements, but activate gene expression through Sp1 sites.10 However, DEC1 does not regulate p53 expression. Instead, DEC1 specifically attenuates p53-dependent transactivation of MIC-1, but not other p53 target genes, including two cell cycle regulators (p21 and GADD45α), four pro-apoptotic genes (Puma, Bax, PolH and FDXR) and Mdm2. Consistent with this, knockdown of DEC1 enhances p53-dependent induction of MIC-1 and apoptosis in response to DNA damage. Moreover, upon knockdown of MIC-1, the effect of DEC1 knockdown on p53-dependent cell death is abrogated. These findings suggest that DEC1 forms a feedback loop with p53 to control the response of DNA damage-induced cell survival vs. cell death via MIC-1. To characterize the mechanism by which DEC1 differentially affects the ability of p53 to regulate its target genes, we found that DEC1 associates with the p53 tetramerization domain through its bHLH motif. Importantly, we noticed that the distance between the DEC1-RE and p53-RE on the MIC-1 promoter is only 17 nt, but more than 280 nt on the p21 and Mdm2 promoters. ChIP and ChIP-reChIP assays showed that DEC1 binds to the MIC-1, p21 and Mdm2 promoters but co-localizes with p53 only on the MIC-1 promoter. In addition, DEC1 preferentially suppresses p53 binding to the MIC-1 promoter upon DNA damage. Thus, we hypothesize that both DEC1 and p53 have to bind to the same promoter in close proximity to allow DEC1-p53 interaction, which is prerequisite for DEC1 to decrease p53 DNA-binding affinity and consequently reduce MIC-1 expression (Fig. 1). This unique regulatory loop between DEC1 and p53 provides a clue for investigating other p53 target genes potentially regulated by DEC1. Figure 1. A model for DEC1 to differentially modulate p53-dependent gene expression. Upon binding to the p53-responsive element (p53-RE), p53 induces an arrary of pro-survival and pro-apoptotic genes, including p21, Mdm2, Bax, MIC-1 and DEC1. ... p53 is frequently mutated in tumors, and most tumor-derived p53 mutations are located in the DNA-binding domain but not the tetramerization domain. Importantly, tumor-derived p53 mutants have gain-of-function activities contributing to tumorigenesis and chemotherapeutical resistance. Since p53 tetramerization domain is found to interact with DEC1, it is likely that DEC1 and mutant p53 physically interact on a promoter that carries DEC1-RE and mutant p53-RE. Thus, it would be of great interest to examine whether DEC1 modulates the function of mutant p53 in tumor progression and the resistance of tumors to therapy. In addition, as a member of the p53 family, p63 and p73 share a high-sequence identity with p53 in the tetramerization domain along with the transactivation and DNA-binding domains. Interestingly, p63 and p73 not only possess p53-like functions in tumor suppression, but also play a key role in development. Thus, it would be interesting to examine whether DEC1 physically interacts with p63 and p73 on a promoter that carries DEC1-RE and p53-RE, and whether DEC1 modulates the function of p63 and p73 in tumor suppression and development.