p53 signaling modulation of cell cycle arrest and viral replication in porcine circovirus type 2 infection cells
- Dan Xu†1,
- Qian Du†1,
- Cong Han1,
- Zengguo Wang1,
- Xiujuan Zhang1,
- Tongtong Wang1,
- Xiaomin Zhao1,
- Yong Huang1Email author and
- Dewen Tong1Email author
© The Author(s) 2016
Received: 12 May 2016
Accepted: 20 September 2016
Published: 29 November 2016
Porcine circovirus type 2 (PCV2) is a ubiquitous pathogen in the swine industry worldwide. Previous studies have shown that PCV2 infection induces host cell apoptosis through up-regulation of p53. To further identify the regulatory roles of p53 signaling in the process of PCV2 infection, we established p53 gene knockout PK15 cell lines using the genomic editor tool CRISPR/Cas9, and further investigated the roles of p53 in modulating the cell cycle and viral replication in this study. The results show that PCV2 infection induced obvious S phase accumulation in wild-type PK15 cells and a compromised S phase accumulation in the p53 gene mutation cells (813PK15 p53m/m ), but did not induce obvious S phase accumulation in the p53 gene knockout cells (148PK15 p53−/−) compared with the respective mock infection. PCV2 infection activated p53 signaling, up-regulated the expression of p21, Cyclin E, and down-regulated Cyclin A, CDK2. In p53 deficient cells, however, PCV2-induced changes in Cyclin A, CDK2, and Cyclin E were efficiently reversed to the basal levels. Detection of PCV2 replication showed decreased viral ORF1 genomic DNA in p53 deficient cells (148PK15 p533−/−) and p53 mutated cells (813PK15 p53m/m ) compared with p53 wild-type cells after different synchronization treatment. Furthermore, PCV2 viral genomic DNA and Cap protein levels were higher in the cells released from S phase synchronized cells than in the cells released from the G0/G1 phase or G2/M phase-synchronized, or asynchronous cells after 18 h post-infection. Taken together, this study demonstrates that PCV2 infection induces S phase accumulation to favor viral replication in host cells through activation of the p53 pathway.
PCV2, belonging to the family Circoviridae, is the main pathogen to cause porcine circovirus associated diseases (PCVAD) , posing a huge threat for world pig husbandry . As a tiny DNA virus, PCV2 infection requires host cells to provide necessary resources for replication themselves, thus disturbing a variety of cell signaling pathways to modulate the host cell cycle, proliferation, survival and death to facilitate their infection and replication [3, 4]. Among the signaling pathways, p53 signaling is essential for control of quiescent cell entry into the cell cycle, and regulating cellular DNA replication . However, the roles of p53 signaling in modulating cell cycle and PCV2 replication has not been defined up to date.
Numbers of studies have broadened our understanding of the roles of p53 signaling in the process of different virus infection and replication. For instance, Kaposi’s sarcoma herpesvirus (KSHV) activates host p53 signal and induces G2 phase arrest to promote the onset of virus replication . Prototype foamy virus (PFV) promotes p53 level increase by knockdown of Pirh2 to contribute to the latency of PFV infection . Herpes simplex virus type 2 infection can phosphorylate p53 protein to induce the G0/G1 phase arrest . PRRSV manipulates the host factors mdm2 and p53 via its Nsp1α to increase viral replication at the early stage of infection . Indeed, previous studies have shown that PCV2 ORF3 protein specifically interacts with porcine ubiquitin E3 ligase Pirh2 to promote p53 accumulation , playing an important role in PCV2 pathogenesis , which indicates the key role of p53 in the interaction of PCV2 and the host. However, in-depth study of the roles of p53 signaling in the process of PCV2 was limited due to lacking of p53 deficient cell line in porcines.
In this study, with the help of the CRISPR/Cas9 system, we constructed p53 deficient and mutant porcine cell lines, and further detected and compared the difference of cell cycle profiles and viral replication between the p53 wild-type, p53 deficient and p53 mutant porcine cell lines. This study allows us to deeply explore and confirm the roles of p53 signaling in modulating cell cycle and PCV2 replication.
Materials and methods
Cells, virus and antibodies
Porcine kidney 15 (PK15) cells purchased from ATCC (CCL-33) were cultured in Dulbecco’s Modified Eagle’s Medium (Gibco BRL, Gaithersburg, MD, USA) supplemented with 10% heat-inactivated fetal bovine serum (Thermo Scientific HyClone, Beijing, China), and incubated at 37 °C in a 5% CO2 atmosphere incubator. The PCV2 strains (GenBank No. EU366323) used in this study were isolated and purified previously by our team and stocked in our laboratory, the UV-inactivation was performed by UV radiation of the virus for 45 min in the hood. The anti-PCV2 Cap primary antibodies were produced by our team [12, 13]. The primary monoclonal rabbit antibodies of p53, p21 and anti-BrdU were purchased from Cell Signaling (Cell Signaling Technology, Danvers, MA, USA). CDK2, Cyclin A and Cyclin E antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, California, CA, USA). The monoclonal antibody of β-actin was purchased from sigma (Sigma-Aldrich, St. Louis, MO, USA). The FITC goat anti-mouse IgG was purchased from BD Biosciences (BD, San Jose, CA, USA).
Cell cycle analysis
The ratio of cells in each phase of the cell cycle was determined by DNA content using propidium iodide (PI) staining followed by flow cytometric analysis. The cells plated at a density of 1 × 106 cells/flask were treated with the indicated Multiplicity of infection (MOI) of PCV2 for the indicated times as described in the figure legends. The cells were trypsinized, washed twice with PBS, and fixed with 70% ice-cold ethanol at −20 °C overnight. Fixed cells were washed with cold PBS and resuspended with PI staining solution containing 50 mg/mL PI (Sigma-Aldrich), 100 mg/mL RNase A (TIANGEN Biotech, Beijing, China), and incubated in the dark for 30 min. The samples were analyzed using a flow cytometer (Accuri™ C6, BD Biosciences, San Diego, CA, USA).
CRISPR/cas9 KO cell
Targeting sites in the p53 gene were selected using the CRISPR program (Genome Engineering. Broad Institute Cambridge, MA, USA) Oligonucleotide pairs for the target sequences were annealed and the resulting fragments were then cloned into the BsmB I sites of lentiCRISPRv2 plasmid (Addgene), and co-transfected into HEK293T cells with the packaging plasmids psPAX2 (AddGene 12260) to generate the lentivirus. 72 h after the transfection, the supernatant was collected after three cycles of frozen-thawed. Titers of the obtained lentivirus expressing the target sequences were determined by qPCR. Finally, the CRISPR/Cas9 mediated P53 knockout cells were selected from lentivirus infected PK15 cell lines that were cultured in puromycin (500 ng/mL) DMEM medium for at least 14 days. Genomic DNA sequence from PK15 cells was determined using primers: 148-F: 5′-GACTCCTGTTGTTCCCATCCA-3′; 148-R: 5′-AGGGAGCCAGCAGTCAAATG-3′; 813-F: 5′-GGGACGGAACAGCTTTGAGGT-3′; 813-R: 5′-CTGTTGGCAAATGCCCCAAA-3′.
Cells synchronized in G1/G0 phase were obtained by serum starvation. PK-15 cells were cultured in serum-free medium for 24 h or 48 h, and then cells were washed with PBS and plated in fresh media to start PCV2 incubation for 1 h and cultured in 2% FBS DMEM medium for 18 or 24 h for later analysis. Double thymidine block was used for early S phase synchronization. The cells were treated for 12 h with 2 mM thymidine, after which cells were washed and released into fresh media with MOI = 1 PCV2 virus then incubated for 1 h, and cultured in 2% FBS DMEM medium for 18 h. The cells were treated with 100 ng/mL nocodazole for 16 h until arrest at the G2/M phase, then the cells were released by washing with PBS and plated in fresh media to start PCV2 incubation for 1 h and culture in 2% FBS DMEM medium for 18 h for later analysis.
Detection of virus replication
The cells were seeded in culture plates at a density of 1 × 106 cells/well, and cultured to reach approximately 60–70% confluence. PCV2 strains were used to infect the cells at a multiplicity of infection of 1. Viral ORF1 fragments were determined in each of the PK15 cell lines using primer PCV-F: 5′-AGTACCGGGAGTGGTAGGAG-3′; PCV-R: 5′-GTTGAATTCTGGCCCTGCTC-3′. The supernatant and the attached cells were collected together to extract the DNA.
BrdU incorporation assay
For labeling of S-phase cells, BrdU was added in mid-log phase cells at a final concentration of 10 µM and incubated for 1 h at 37 °C. The cells were harvested and washed with PBS + 1% BSA. The cells were further resuspended and fixed overnight in chilled 70% ethanol at a cell density of 2 × 106 cells/mL at 4 °C. Further, ethanol was removed and the cells were incubated in 2 N HCL+ 0.5% Triton X-100 solution for 30 min at RT followed by washing in 0.1 M Sodium tetraborate solution for 2 min. Then the cells were resuspended in PBS/0.5% Tween-20 + 1% BSA and incubated with anti-BrdU antibody for 1 h at RT. The cells were washed again with the same buffer and incubated in PBS/0.5% Tween-20 + 1% BSA with FITC goat antimouse antibody for 0.5 h at RT. Then the cells were washed and resuspended in RNase + PI at RT for 0.5 h min and finally centrifuged. The cells were collected for flow cytometer analysis.
The protein expression was measured by western blot as described previously . Briefly, PK15 cells were cultured in a flask, infection was as indicated at an MOI of PCV2 virus and harvested at an 18 h interval as described above. Whole-cell protein extract was prepared as described above. The protein extract from each group was loaded to 12% SDS-PAGE, and transferred to PVDF membranes. After blocking for 1 h at room temperature, the membranes were incubated with primary antibody at 4 °C overnight. After being incubated for 1.5 h with HRP-conjugated IgG second monoclonal antibody at room temperature, the membranes were immersed in an ECL reagent (Pierce, Rorkford, IL, USA) and visualized in X-ray films. The optical density of each band was quantified using ImageJ analysis software (NIH, NY, USA) with β-actin as an internal control.
All data were presented as “mean ± SEM” in triplicate, and analyzed with the Student’s t test using SPSS 17.0 software (SPSS Inc., Chicago, IL, USA). Differences with the controls were considered significant when p < 0.05.
PCV2-infected cells accumulated at the S phase of the cell cycle
We further addressed serum-starved quiescence cells to monitor the quantification of DNA cycle fractions upon PCV2 infection. PK15 cells were serum-starved for 24 h followed by PCV2 infection. S phase cell populations increased by 1.32-, 1.46- and 1.31-fold at 12, 24 and 48 h pi, respectively, in PCV2-infected cells, compared to that in mock infection cells, which was accompanied by a concomitant decrease in the G0/G1 or G2/M phase cell populations (Figure 1B), indicating that PCV2 blocked the cell cycle at the S phase. At 48 h pi, the G2/M population of the cell was markedly reduced, suggesting the complete blockage of the S-G2 transition in PCV2-infected cells. Meanwhile, viral ORF1 genomic DNA in PCV2-infected cells (MOI = 1) was monitored by qPCR from the asynchronic and synchronic of infection cells, the total ORF1 genomic DNA were amplified from 12 h, and significantly increased at 24 and 48 h pi (Figure 1C). To determine whether PCV2-induced S phase accumulation was dependent on the viral replication, 1 MOI of UV-inactivated PCV2 was inoculated with synchronized PK-15 for 24 h to measure the cell cycle profiles by flow cytometry. The results show that there was no significant difference for cell cycle distribution between the UV-inactivated PCV2 group and the mock infection group (Figure 1D), suggesting that UV-inactive PCV2 virus did not alter cell cycle distribution, and viral replication was required for induction of cell cycle arrest in PCV2-infected cells. These results indicate that PCV2 infection induces numerous host cells to accumulate in the S phase, which might be beneficial for virus replication.
Loss of p53 impairs PCV2 induced S phase accumulation and viral propagation
p53 mediates up-regulation of p21 and Cyclin E and down-regulation of Cyclin A, CDK 2 in PCV2-infected cells
However, in the p53 knockout PK15 cells (148PK15 P53−/−), p53 could not be detected, and p21 also could not be detected at 18 and 36 h after PCV2 infection. In accordance with these changes, the p53 knockout cells exhibited relatively higher levels of Cyclin A and CDK2 and relatively lower levels of Cyclin E compared with wild-type cells at 18 and 36 h pi (Figure 4B). Meanwhile, p53 mutated PK15 cells (813PK15 P53m/m ) showed relatively lower levels of p53, Cyclin E and relatively higher levels of Cyclin A when compared with WT cells, but relatively lower levels of Cyclin A and CDK2 when compared with 148PK15 P53−/− cells at 18 and 36 h pi. These results suggest that PCV2 induced S phase accumulation through p53 mediated up-regulation of p21, Cyclin E and down-regulation of Cyclin A, CDK 2 proteins in PK15 cells.
S phase accumulation promotes PCV2 replication
Real-time PCR analysis of the genomic DNA of PCV2 shows that the replication levels of ORF1 DNA were higher in the WT cells released from G1/S-synchronized cells than those in the WT cells released from G0/G1 phase or G2/M phase synchronized, or asynchronous WT cells (Figure 5D). In addition, the levels of the PCV2 Cap protein also were higher in the WT cells released from G1/S-synchronized cells than those in the WT cells released from G0/G1 phase or G2/M phase synchronized or asynchronous cells (Figure 5E). However, the viral DNA levels and Cap protein levels were lower in the 148PK15 P53−/− and 813PK15 P53m/m cells released from the G1/S phase synchronized cells than those in the WT cells released from the G1/S phase synchronized cells (Figure 5D and F). These results suggest that PCV2 induction of S phase accumulation via p53 signaling might be beneficial for virus replication.
Viruses evolve complex strategies to override cell cycle checkpoints forcing host cells into specific phases to support viral replication. Some viruses interfere with the p53 surveillance pathways to promote replication [16, 17], other viruses exhibited a p53-independent way for its replication [18, 19]. In this study, we focused on whether the p53 signaling pathway contributes to PCV2 induced cell cycle progression and virus replication. The results show that PCV2 infection activated the p53 signaling to up-regulate p21 and Cyclin E while down-regulating Cyclin A and CDK2, forcing the infected cells to stay in the replicative S phase. The roles of p53 in PCV2 replication were further tested by infection of p53 knockout and p53 mutant cells, in which the progression of the cell cycle was less effected by PCV2 and the replication of virus appears a relative lower level.
In this study, the first question we answered is whether PCV2 overrides cell cycle checkpoints for its benefits. In both asynchronic and synchronic cells, PCV2 induced cell cycle arrest at the S phase and promoted both PCV2 capsid protein and PCV2 progeny production, whereas UV-inactivated PCV2 did not exhibit S-phase accumulation. Besides this, PCV2 genomic DNA and Cap protein levels were higher in the WT cells released from G1/S-synchronized cells than those in the WT cells released from G0/G1 phase or G2/M phase synchronized, or asynchronous WT cells, suggesting that S phase accumulation might be beneficial for virus replication. Our data explain and agree with the previous reports that PCV2 DNA synthesis dramatically increases in PK15 cells right before mitosis , while the cell cycle regulator protein Cyclin A overexpression suppressed PCV2 replication . Thus, we confirmed that PCV2 infection induced S-phase accumulation for its multiplication.
The other main question we answered is the roles of p53 signaling in modulating cell cycle arrest and PCV2 replication. The function of p53 in regulating the cell cycle progress of PCV2-infected cells was explored by comparing p53 wild-type, knockout and mutant cell lines. Following PCV2 infection, p53 knockout caused significant reduction of DNA synthesis compared with the wild-type cell, whereas the p53 knockout cells exhibited a relatively stabilized high DNA synthesis rate whether in the presence or absence of PCV2 in the BrdU assay. We also observed that PCV2 genomic DNA levels and Cap protein levels were lower in the 148PK15 P53−/− and 813PK15 P53m/m cells released from G1/S phase synchronized cells than those in the WT cells released from G1/S phase synchronized cells. These results suggest that p53 plays a pivotal role in inducing S-phase cell accumulation and viral replication in PCV2-infected cells.
Actually, the role of p53 interplays between the host and virus has been extensively studied. Some viruses hijack the p53 apoptotic pathway to facilitate virus invasion, like Influenza A virus , Reovirus , and Epstein–Barr virus  while other virus proteins directly interact with p53 effecting both p53 DNA-binding affinity and transcriptional ability, such as Hepatitis C virus , and Hepatitis B virus . In addition, some viruses specifically interrupt the cellular p53–p21 pathway to modulate the host cell cycle, blocking cellular DNA synthesis in generating viral nucleotide pools, like Respiratory syncytial virus , Herpes simplex virus type 2 (HSV-2), and Simian virus 40 (SV40). However, unlike HSV-2, which increases p21 protein levels by phosphorylation of the p53 protein at Ser20 , SV40 induced p53 phosphorylation is accompanied by Ser15 , as SV40 activating the ATR-Delta p53 signaling to maintain S phase environment, and to manipulate polymerases . This may explain our data that in the CRISPR/Cas 9 mediated 148PK15 P53−/− cells, PCV2 infection did not induce S phase accumulation, resulting in a relative lower virus production, while the 813PK15 P53m/m cells that mutated original S271 and G272 sites of porcine p53 into R271 showed a slightly weakened function of p53 signaling undergoing PCV2 infection, resulting in a slight relative decreased S phase accumulation compared with the wild-type cell. Single amino acid mutation of p53 exhibited a different function, the human R273 site mutation induced a resistance to drug which induced apoptosis, while the mouse R270 residue is subject to stress-induced modifications [29, 30]. Similarly, in this study, the deletion of a single amino acid located at the sensitive key site, affected the progression of cell cycle in 813PK15 P53m/m cells. Interestingly, the delta p53 signaling, an alternative p53 isoform deleted from 256 to 322 amino acid residues, is activated upon the DNA virus SV40 infection or when the host cells encounter a single-strand DNA break, resulting in up-regulation of p21 and down-regulation of Cyclin A-CDK2 in S phase [17, 31]. In the current study, following PCV2 infection, an increase of p21 and a decrease of Cyclin A-CDK2 was observed, while in the p53 deficient cells, the level of expression of Cyclin A-CDK2 was efficiently altered to the basal level.
The CRISPR/Cas9-mediated gene editing system has emerged as a powerful and efficient tool to manipulate the genomes of potential targets to understand pathogeneses [32, 33]. Taking advantage of this technology allowed us to conduct basic research on p53 signaling in porcine cell lines. Even though the exact molecular mechanism of PCV2 infected cell response to p53 signaling still needs to be further studied, our data presented here confirmed that PCV2 induced cell cycle accumulates at the S phase for its replication, and p53 signaling contributes to this modulation process.
porcine circovirus type 2
porcine circovirus associated diseases
Kaposi’s sarcoma herpesvirus
prototype foamy virus
porcine kidney 15
multiplicity of infection
Herpes simplex virus type 2
Simian virus 40
The authors declare that they have no competing interests.
DX and YH designed the experiments, interpreted the data and wrote the article. DX performed the experiments with assistance and advice from QD, CH, ZW, XZ, TW, XZ. YH and DT revised the paper. All authors read and approved the final manuscript.
This work was supported by the National Natural Science Foundation of China (Grant No. 31372401, 31372411), Key Project of Shaanxi Province Science and Technology Innovation Team (2013 KCT-28), the innovation project of science and technology plan projects in Shaanxi province (2016KTCL02-13), and the Fundamental Research Funds for the Central Universities (2452015155, 2452016116).
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- Opriessnig T, Meng XJ, Halbur PG (2007) Porcine circovirus type 2 associated disease: update on current terminology, clinical manifestations, pathogenesis, diagnosis, and intervention strategies. J Vet Diagn Invest 19:591–615View ArticlePubMedGoogle Scholar
- Allan GM, Kennedy S, McNeilly F, Foster JC, Ellis JA, Krakowka SJ, Meehan BM, Adair BM (1999) Experimental reproduction of severe wasting disease by co-infection of pigs with porcine circovirus and porcine parvovirus. J Comp Pathol 121:1–11View ArticlePubMedGoogle Scholar
- Wei L, Zhu Z, Wang J, Liu J (2009) JNK and p38 mitogen-activated protein kinase pathways contribute to porcine circovirus type 2 infection. J Virol 83:6039–6047View ArticlePubMedPubMed CentralGoogle Scholar
- Wei L, Liu J (2009) Porcine circovirus type 2 replication is impaired by inhibition of the extracellular signal-regulated kinase (ERK) signaling pathway. Virology 386:203–209View ArticlePubMedGoogle Scholar
- Karimian A, Ahmadi Y, Yousefi B (2016) Multiple functions of p21 in cell cycle, apoptosis and transcriptional regulation after DNA damage. DNA Repair 42:63–71View ArticlePubMedGoogle Scholar
- Balistreri G, Viiliainen J, Turunen M, Diaz R, Lyly L, Pekkonen P, Rantala J, Ojala K, Sarek G, Teesalu M, Denisova O, Peltonen K, Julkunen I, Varjosalo M, Kainov D, Kallioniemi O, Laiho M, Taipale J, Hautaniemi S, Ojala PM (2016) Oncogenic herpesvirus utilizes stress-induced cell cycle checkpoints for efficient lytic replication. PLoS Pathog 12:e1005424View ArticlePubMedPubMed CentralGoogle Scholar
- Dong LL, Cheng QQ, Wang ZH, Yuan PP, Li Z, Sun Y, Han S, Yin J, Peng BW, He XH, Liu WH (2015) Human Pirh2 is a novel inhibitor of prototype foamy virus replication. Viruses 7:1668–1684View ArticlePubMedPubMed CentralGoogle Scholar
- Zhou Q, Zhu M, Zhang H, Yi T, Klena JD, Peng Y (2012) Disruption of the p53–p21 pathway inhibits efficiency of the lytic-replication cycle of herpes simplex virus type 2 (HSV-2). Virus Res 169:91–97View ArticlePubMedGoogle Scholar
- Wang X, Shao C, Wang L, Li Q, Song H, Fang W (2016) The viral non-structural protein 1 alpha (Nsp1alpha) inhibits p53 apoptosis activity by increasing murine double minute 2 (mdm2) expression in porcine reproductive and respiratory syndrome virus (PRRSV) early-infected cells. Vet Microbiol 184:73–79View ArticlePubMedGoogle Scholar
- Karuppannan AK, Liu S, Jia Q, Selvaraj M, Kwang J (2010) Porcine circovirus type 2 ORF3 protein competes with P53 in binding to Pirh2 and mediates the deregulation of P53 homeostasis. Virology 398:1–11View ArticlePubMedGoogle Scholar
- Liu J, Chen I, Du Q, Chua H, Kwang J (2006) The ORF3 protein of porcine circovirus type 2 is involved in viral pathogenesis in vivo. J Virol 80:5065–5073View ArticlePubMedPubMed CentralGoogle Scholar
- Huang Y, Xing N, Wang Z, Zhang X, Zhao X, Du Q, Chang L, Tong D (2015) Ultrasensitive detection of RNA and DNA viruses simultaneously using duplex UNDP-PCR assay. PLoS One 10:e0141545View ArticlePubMedPubMed CentralGoogle Scholar
- Du Q, Huang Y, Wang T, Zhang X, Chen Y, Cui B, Li D, Zhao X, Zhang W, Chang L, Tong D (2016) Porcine circovirus type 2 activates PI3K/Akt and p38 MAPK pathways to promote interleukin-10 production in macrophages via Cap interaction of gC1qR. Oncotarget 7:17492–17507PubMedPubMed CentralGoogle Scholar
- Ding L, Huang Y, Dai ML, Zhao XM, Du Q, Dong F, Wang LL, Huo RC, Zhang WL, Xu XG, Tong DW (2013) Transmissible gastroenteritis virus infection induces cell cycle arrest at S and G2/M phases via p53-dependent pathway. Virus Res 178:241–251View ArticlePubMedGoogle Scholar
- Poon B, Grovit-Ferbas K, Stewart SA, Chen IS (1998) Cell cycle arrest by Vpr in HIV-1 virions and insensitivity to antiretroviral agents. Science 281:266–269View ArticlePubMedGoogle Scholar
- Luo Y, Chen AY, Qiu J (2011) Bocavirus infection induces a DNA damage response that facilitates viral DNA replication and mediates cell death. J Virol 85:133–145View ArticlePubMedGoogle Scholar
- Rohaly G, Korf K, Dehde S, Dornreiter I (2010) Simian virus 40 activates ATR-Delta p53 signaling to override cell cycle and DNA replication control. J Virol 84:10727–10747View ArticlePubMedPubMed CentralGoogle Scholar
- Dahl J, You J, Benjamin TL (2005) Induction and utilization of an ATM signaling pathway by polyomavirus. J Virol 79:13007–13017View ArticlePubMedPubMed CentralGoogle Scholar
- Kucharski TJ, Gamache I, Gjoerup O, Teodoro JG (2011) DNA damage response signaling triggers nuclear localization of the chicken anemia virus protein Apoptin. J Virol 85:12638–12649View ArticlePubMedPubMed CentralGoogle Scholar
- Tischer I, Peters D, Rasch R, Pociuli S (1987) Replication of porcine circovirus: induction by glucosamine and cell cycle dependence. Arch Virol 96:39–57View ArticlePubMedGoogle Scholar
- Tang Q, Li S, Zhang H, Wei Y, Wu H, Liu J, Wang Y, Liu D, Zhang Z, Liu C (2013) Correlation of the cyclin A expression level with porcine circovirus type 2 propagation efficiency. Arch Virol 158:2553–2560View ArticlePubMedGoogle Scholar
- Nailwal H, Sharma S, Mayank AK, Lal SK (2015) The nucleoprotein of influenza A virus induces p53 signaling and apoptosis via attenuation of host ubiquitin ligase RNF43. Cell Death Dis 6:e1768View ArticlePubMedPubMed CentralGoogle Scholar
- Pan D, Pan LZ, Hill R, Marcato P, Shmulevitz M, Vassilev LT, Lee PW (2011) Stabilisation of p53 enhances reovirus-induced apoptosis and virus spread through p53-dependent NF-kappaB activation. Br J Cancer 105:1012–1022View ArticlePubMedPubMed CentralGoogle Scholar
- Al-Salam S, Awwad A, Sudhadevi M, Daoud S, Nagelkerke NJ, Castella A, Chong SM, Alashari M (2013) Epstein–Barr virus infection correlates with the expression of COX-2, p16(INK4A) and p53 in classic Hodgkin lymphoma. Int J Clin Exp Pathol 6:2765–2777PubMedPubMed CentralGoogle Scholar
- Otsuka M, Kato N, Lan K, Yoshida H, Kato J, Goto T, Shiratori Y, Omata M (2000) Hepatitis C virus core protein enhances p53 function through augmentation of DNA binding affinity and transcriptional ability. J Biol Chem 275:34122–34130View ArticlePubMedGoogle Scholar
- Chan C, Wang Y, Chow PK, Chung AY, Ooi LL, Lee CG (2013) Altered binding site selection of p53 transcription cassettes by hepatitis B virus X protein. Mol Cell Biol 33:485–497View ArticlePubMedPubMed CentralGoogle Scholar
- Bian T, Gibbs JD, Orvell C, Imani F (2012) Respiratory syncytial virus matrix protein induces lung epithelial cell cycle arrest through a p53 dependent pathway. PLoS One 7:e38052View ArticlePubMedPubMed CentralGoogle Scholar
- Rohaly G, Chemnitz J, Dehde S, Nunez AM, Heukeshoven J, Deppert W, Dornreiter I (2005) A novel human p53 isoform is an essential element of the ATR-intra-S phase checkpoint. Cell 122:21–32View ArticlePubMedGoogle Scholar
- Rieckmann T, Kriegs M, Nitsch L, Hoffer K, Rohaly G, Kocher S, Petersen C, Dikomey E, Dornreiter I, Dahm-Daphi J (2013) p53 modulates homologous recombination at I-SceI-induced double-strand breaks through cell-cycle regulation. Oncogene 32:968–975View ArticlePubMedGoogle Scholar
- Wong RPC, Tsang WP, Chau PY, Co NN, Tsang TY, Kwok TT (2007) p53-R273H gains new function in induction of drug resistance through down-regulation of procaspase-3. Mol Cancer Ther 6:1054–1061View ArticlePubMedGoogle Scholar
- Kocher S, Rieckmann T, Rohaly G, Mansour WY, Dikomey E, Dornreiter I, Dahm-Daphi J (2012) Radiation-induced double-strand breaks require ATM but not Artemis for homologous recombination during S-phase. Nucleic Acids Res 40:8336–8347View ArticlePubMedPubMed CentralGoogle Scholar
- Liu T, Shen JK, Li ZH, Choy E, Hornicek FJ, Duan ZF (2016) Development and potential applications of CRISPR-Cas9 genome editing technology in sarcoma. Cancer Lett 373:109–118View ArticlePubMedGoogle Scholar
- Mei Y, Wang Y, Chen HQ, Sun ZS, Ju XD (2016) Recent progress in CRISPR/Cas9 technology. J Genet Genom 43:63–75View ArticleGoogle Scholar