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Background: The Alternative Lengthening of Telomeres (ALT) pathway represents a non-canonical mechanism of telomere maintenance that operates independently of the conventional telomerase activity. The three biologically significant proteins, designated as SMARCAL1 (SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A-like protein 1), DAXX (Death domain-associated protein 6) and ATRX (alpha-thalassemia/mental retardation, X-linked) are associated with ALT in certain cancer types. The purpose of this study was to identify the most high-risk nsSNPs (non-synonymous Single Nucleotide Polymorphisms) within these three genes and assess their impacts on the structure and function of the proteins they encode. Methods: The reported genetic polymorphisms of SMARCAL1, DAXX and ATRX genes were retrieved from the Ensembl database. Later, various computational tools like PROVEAN, PolyPhen2, SNPs and GO, SNAP2, Predict-SNP, Panther and PMut were used to predict the most deleterious nsSNPs. MutPred was used to understand the underlying molecular reasons of those nsSNPs being deleterious, followed by prediction of Post Translational Modification Sites (PTMs) using ModPred. I-Mutant and MUpro were used to predict the effect of SNP on energy stability. Later, 3D clustering analysis was done using Mutation 3D server. Moreover, ConSurf was utilized to identify the conservation scores of wild-type amino acids. Additionally, the NCBI conserved domain search tool was employed to pinpoint conserved domains within these three proteins. Project-Hope helped for biophysical validation, followed by prediction of these genes’ interaction and function by using GeneMANIA. Result: Analysis on SMARCAL1 protein revealed that among 665 nsSNPs, four were identified as the most deleterious: L578S, T581S, P582A, and P582S. Similarly, within the DAXX protein, among a pool of 480 nsSNPs, P284S, R230C, and R230S were found out to be the most deleterious variants. In case of ATRX protein, V178D, R246C, and V277G, from the total of 1009 nsSNPs, were predicted to be the most deleterious. All these nsSNPs were found to occur at residue positions that are 100 % conserved within protein domains and were predicted to be most damaging from both structural and functional perspectives and highly destabilizing to their corresponding proteins. Conclusion: Computational investigation on the 3 proteins-SMARCAL1, DAXX and ATRX through different bioinformatics analysis tools concludes that the identified high risk nsSNPs of these proteins are pathogenic SNPs. These variants potentially exert functional and structural influences, thus making them valuable candidates for future genetic studies.

Uveal melanomas (UMs) represent rare malignant tumors associated with grim prognosis for the majority of patients. DAXX (Death Domain-Associated Protein), HJURP (Holliday Junction Recognition Protein) and CENPA (Centromere Protein A) proteins are implicated in epigenetic mechanisms, now in the spotlight of cancer research to better understand the molecular background of tumorigenesis. Herein, we investigated their expression in UM tissues using immunohistochemistry and explored possible correlations with a multitude of clinicopathological and survival parameters. The Cancer Genome Atlas Program (TCGA) was used for the investigation of their mRNA levels in UM cases. Nuclear DAXX expression correlated with an advanced T-stage (p = 0.004), while cytoplasmic expression marginally with decreased disease-free survival (DFS) (p = 0.084). HJURP nuclear positivity also correlated with advanced T-status (p = 0.054), chromosome 3 loss (p = 0.042) and increased tumor size (p = 0.03). More importantly, both nuclear and cytoplasmic HJURP immunopositivity correlated with decreased overall survival (OS) (p = 0.011 and 0.072, respectively) and worse DFS (p = 0.071 and 0.019, respectively). Lastly, nuclear CENPA overexpression was correlated with presence of irido-corneal angle involvement (p = 0.015) and loss of chromosome 3 (p = 0.041). Nuclear and cytoplasmic CENPA immunopositivity associated with decreased OS (p = 0.028) and DFS (p = 0.018), respectively. HJURP and CENPA mRNA overexpression exhibited strong association with tumor epithelioid histology and was linked to worse prognosis. Our results show the compounding role of DAXX, HJURP and CENPA in UM carcinogenesis, designating them as potential biomarkers for assessing prognosis and possible targets for novel therapeutic interventions.

Abstract Promyelocytic leukemia nuclear bodies (PM NBs), often referred to as membraneless organelles, are dynamic macromolecular protein complexes composed of a PML protein core and other transient or permanent components. PML NBs have been shown to play a role in a wide variety of cellular processes. This review describes in detail the diverse and complex interactions between small and medium size DNA viruses and PML NBs that have been described to date. The PML NB components that interact with small and medium size DNA viruses include PML protein isoforms, ATRX/Daxx, Sp100, Sp110, HP1, and p53, among others. Interaction between viruses and components of these NBs can result in different outcomes, such as influencing viral genome expression and/or replication or impacting IFN-mediated or apoptotic cell responses to viral infection. We discuss how PML NB components abrogate the ability of adenoviruses or Hepatitis B virus to transcribe and/or replicate their genomes and how papillomaviruses use PML NBs and their components to promote their propagation. Interactions between polyomaviruses and PML NBs that are poorly understood but nevertheless suggest that the NBs can serve as scaffolds for viral replication or assembly are also presented. Furthermore, complex interactions between the HBx protein of hepadnaviruses and several PML NBs-associated proteins are also described. Finally, current but scarce information regarding the interactions of VP3/apoptin of the avian anellovirus with PML NBs is provided. Despite the considerable number of studies that have investigated the functions of the PML NBs in the context of viral infection, gaps in our understanding of the fine interactions between viruses and the very dynamic PML NBs remain. The complexity of the bodies is undoubtedly a great challenge that needs to be further addressed.

Genome sequencing has revealed the importance of epigenetic regulators in tumorigenesis. The genes encoding the chromatin remodeling complex DAXX:ATRX are frequently mutated in pancreatic neuroendocrine tumors; however, the underlying mechanisms of how mutations contribute to tumorigenesis are only partially understood, in part because of the lack of relevant preclinical models. Here, we used genetically engineered mouse models combined with environmental stress to evaluate the tumor suppressor functions of Daxx and Atrx in the mouse pancreas. Daxx or Atrx loss, alone or in combination with Men1 loss, did not drive or accelerate pancreatic neuroendocrine tumorigenesis. Moreover, Daxx loss did not cooperate with environmental stresses (ionizing radiation or pancreatitis) or with the loss of other tumor suppressors (Pten or p53) to promote pancreatic neuroendocrine tumorigenesis. However, owing to promiscuity of the Cre promoter used, hepatocellular carcinomas and osteosarcomas were observed in some instances. Overall, our findings suggest that Daxx and Atrx are not robust tumor suppressors in the endocrine pancreas of mice and indicate that the context of a human genome is essential for tumorigenesis. This article has an associated First Person interview with the first author of the paper.

Phenotypic switching is the main cause of the abnormal proliferation and migration of vascular smooth muscle cells (VSMCs). We previously showed that Daxx exerted negative regulatory effect on AngII-induced VSMC proliferation and migration. However, the function of Daxx in VSMC phenotype switching remained unknown. Nicotinate-curcumin (NC) is an esterification derivative of niacin and curcumin that can prevent the formation of atherosclerosis. We found that NC significantly decreased AngII-induced VSMC phenotype switching. Furthermore, NC significantly inhibited AngII-induced cell proliferation and migration. Moreover, NC upregulated Daxx expression and regulated the PTEN/Akt signaling pathway. We concluded that NC inhibited AngII-induced VSMC phenotype switching by regulating the PTEN/Akt pathway, and through a mechanism that might be associated with the upregulation of Daxx expression.

Abstract Background Telomere stability is one of the hallmarks of cancer that promotes cellular longevity, the accumulation of genetic alterations, and tumorigenesis. The loss of death domain-associated protein (DAXX) and α-thalassemia/mental retardation X-linked protein (ATRX) plays a role in telomere lengthening and stability. This study aims to evaluate the prognostic significance of telomere length (TL) and its association with DAXX and ATRX proteins in breast cancer (BC). Our study used the FISH technique to detect peptide nucleic acid (PNA) in the peripheral blood cells of a cohort of BC patients (n = 220) and a control group of apparently healthy individuals (n = 100). Expression of DAXX and ATRX proteins was evaluated using immunohistochemistry (IHC) in all BC tissues. Results Patients with a shorter TL had worse disease-free survival (DFS) and overall survival (OS). There were significant associations between shorter TL and advanced disease stages, lymph node metastasis, and positive HER2/neu expression. DAXX protein expression was significantly correlated with TL. Lower DAXX expression was significantly with shorter DFS. Conclusion Assessing TL can be used as a worthy prognostic indicator in BC patients. Specifically, short TL had a poor impact on the prognosis of BC patients. Low DAXX expression is associated with poor outcomes in BC. Further mechanistic studies are warranted to reveal the underlying mechanisms of these associations.

BackgroundPancreatic neuroendocrine tumors (PanNETs) are a heterogeneous group of neoplasms with increasing incidence and unpredictable behavior. Whole-exome sequencing recently has shown very frequent somatic mutations in the alpha-thalassemia/mental retardation X-linked (ATRX) and death domain-associated protein (DAXX) genes in PanNETs. And the prognostic significance of altered ATRX/DAXX genes in PanNETs patients have been revealed in several reports. However, many of these include small sample size and hold controversial opinions. To increase statistical power, we performed a systematic review and meta-analysis to determine a pooled conclusion. We examined the impact of altered ATRX/DAXX genes mainly on overall survival (OS), disease-free survival (DFS) and relapse-free survival (RFS) in PanNETs.MethodsEligible studies were identified and quality was assessed using multiple search strategies (last search May 2021). Data were collected from studies about prognostic significance of altered ATRX/DAXX in PanNETs. Studies were pooled, and combined hazard ratios (HRs) with 95% confidence intervals (CIs) were used to estimate strength of the associations.ResultsFourteen studies involving 2313 patients treated for PanNETs were included. After evaluating for publication bias, disease-free survival and relapse-free survival was significantly shortened in patients with altered ATRX/DAXX gene, with combined HR 5.05 (95% confidence interval (CI): 1.58-16.20, P = 0.01) and 3.21 (95% confidence interval (CI): 1.44-7.16, P < 0.01) respectively. However, the combined data showed there were no difference between patients with altered ATRX/DAXX gene or not in overall survival, with a combined HR 0.71 (95% confidence interval (CI): 0.44-1.15, P = 0.23). We also performed a subgroup analysis with metastatic patients in overall survival, showing a combined HR 0.22 (95% confidence interval (CI): 0.11-0.48, P = 0.96). The small number of studies and paucity of multivariate analyses are the limitations of our study.ConclusionsThis is the first rigorous pooled analysis assessing ATRX/DAXX mutation as prognostic biomarkers in PanNETs. Patients with altered ATRX/DAXX gene would have poor DFS according to the combined data. And altered ATRX/DAXX genes in metastatic patients showed a trend towards improved overall survival, although the difference did not reach statistical significance.

Background: Recent advances demonstrate the role of chromatin regulators, including histone variants and histone chaperones, in cancer initiation and progression. Methods: Histone H3K4me3, histone variant centromere protein (CENP-A) and histone chaperones Holliday junction recognition protein (HJURP) as well as DAXX expression were examined immunohistochemically in 95 thymic epithelial tumor (TET) specimens. Our results were compared with the expression profile of DAXX, HJURP and CENP-A in gene expression profiling interactive analysis (GEPIA2). Results: The lymphocyte-poor B3- and C-type TETs were more frequently DAXX negative (p = 0.043). B3 and C-Type TETs showed higher cytoplasmic and nuclear CENP-A (p = 0.007 and p = 0.002) and higher cytoplasmic HJURP H-score (p < 0.001). Higher nuclear CENP-A and cytoplasmic HJURP expression was associated with advanced Masaoka–Koga stage (p = 0.048 and p < 0.001). A positive correlation between HJURP and CENP-A was also observed. The presence of cytoplasmic CENP-A expression was correlated with a favorable overall survival (p = 0.03). CENP-A overexpression in survival analysis of TCGA TETs showed similar results. H3K4me3 expression was not associated with any clinicopathological parameters. Conclusions: Our results suggest a significant interaction between CENP-A and HJURP in TETs. Moreover, we confirmed the presence of a cytoplasmic CENP-A immunolocalization, suggesting also a possible favorable prognostic value of this specific immunostaining pattern.

Backgroud and Purpose Hyperphosphatemia, which is a high inorganic phosphate (Pi) level in the serum, promotes endothelial cells dysfunction and is associated with cardiovascular diseases in patients with chronic kidney diseases (CKD). However, the underlying mechanism of high Pi-induced endothelia cell apoptosis remains unclear. Methods Human umbilical vein endothelial cells (HUVECs) were treated with normal Pi (1.0 mM) and high Pi (3.0 mM), and then cell apoptosis, abnormal gene expression and potential signaling pathway involvement in simulated hyperphosphatemia were examined using flow cytometry, quantitative PCR (qPCR) and western blot analysis. A two-step 5/6 nephrectomy was carried out to induce CKD and biochemical measurements were taken. Results The rat model of CKD revealed that hyperphosphatemia is correlated with an increased death-domain associated protein (DAXX) expression in endothelial cells. In vitro, high Pi increased the mRNA and protein expression level of DAXX in HUVECs, effects that were reversed by additional phosphonoformic acid treatment. Functionally, high Pi resulted in a significantly increased apoptosis in HUVECs, whereas DAXX knockdown markedly repressed high Pi-induced cell apoptosis, indicating that DAXX mediated high Pi-induced endothelial cell apoptosis. High Pi treatment and DAXX overexpression induced the activation of extracellular regulated protein kinases (ERKs), while DAXX knockdown inhibited high Pi-induced ERKs activation. Finally, we demonstrated that DAXX overexpression induced HUVECs apoptosis in the presence of normal Pi, whereas additional treatment with U0126 (a specific ERK inhibitor) reversed that effect. Conclusion Upregulated DAXX promoted high Pi-induced HUVECs apoptosis by activating ERK signaling and indicated that the DAXX/ERK signaling axis may be served as a potential target for CKD therapy.

Abstract Background The death-domain-associated protein (DAXX) was originally identified as a protein that binds to the transmembrane death receptor FAS and enhances both FAS-induced and transforming growth factor-β-dependent apoptosis. In a previous study, we found that nude mice injected with DAXX-overexpressing cells (ES-2-DAXX) accumulated large concentrations of first-generation ascites cells (I ascites cells). The role of DAXX in the development of ascites is unknown. The aim of this study was to analyze the effect of DAXX on proliferation and migration of ascites cells in ovarian cancer in vitro and in vivo. Methods Nude mice were housed in cages with a 14:10 h light:dark cycle; water and food were provided ad libitum. ES-2-DAXX cells (1×106) were injected intraperitoneally into athymic nude mice (8-week-old female mice). After 4 weeks, I ascites cells were collected. The I ascites cells were injected intraperitoneally into athymic nude mice (8-week-old female mice). After 4 weeks, II ascites cells were collected and cultured. Ascites cell survival, migration, and colony formation were measured using colony formation and cell growth assays. Immunofluorescent staining revealed the co-localization of DAXX and promyelocytic leukemia protein (PML) in ascites cell nuclei. Western blotting and immunohistochemistry showed that extracellular signal-related kinase (p-ERK) 1/2 and CEBP-β were highly expressed in tumor tissues formed by II ascites cells. Through immunoprecipitation, we also found that DAXX can interact with CEBP-β. Results DAXX enhanced ascites cell survival, migration, and colony formation. DAXX and PML nuclear foci dramatically increased in a passage-dependent manner in ascites cells, DAXX promoted the tumor growth of ascites cells in vivo, increased ascites cell proliferation in vivo, and enhanced ascites cell survival and migration by activating the ERK signalling pathway and integrating with CEBP-β. Conclusions DAXX can interact with CEBP-β. DAXX can induce ovarian cancer ascites formation by activating the ERK signal pathway and binding to CEBP-β.

Chondrosarcoma is the second most common malignant bone tumor. Current therapies remain ineffective, resulting in poor prognoses. Biomarkers for chondrosarcoma and predictors of its prognosis have not been established. Mental health–related proteins have been associated with the pathogenesis, progression, and prognosis of many cancers, but their association with chondrosarcoma has not been reported. In this study, the expression and clinicopathological significance of the mental health–related proteins DAXX, DRD3, and DISC1 in chondrosarcoma tissue samples were examined, over an 84-months follow-up period. In immunohistochemical analysis, the rates of positive DAXX, DRD3, and DISC1 expression were significantly higher in chondrosarcoma than in osteochondroma tissue (P < 0.01). The percentages of positive DAXX, DRD3, and DISC1 expression were significantly lower in tissues with good differentiation (P < 0.01), AJCC stage I/ II (P < 0.01), Enneking stage I (P < 0.01), and non-metastasis (P < 0.05), respectively. In Kaplan–Meier survival analysis, significantly shorter mean survival times were associated with moderate and poor differentiation (P = 0.000), AJCC stage III/IV (P = 0.000), Enneking stage II/III (P = 0.000), metastasis (P = 0.019), invasion (P = 0.013), and positive DAXX (P = 0.012), and/or DRD3 (P = 0.018) expression. In Cox regression analysis, moderate and poor differentiation (P = 0.006), AJCC stage III/IV (P = 0.013), Enneking stage II/III (P = 0.016), metastasis (P = 0.033), invasion (P = 0.011), and positive DAXX (P = 0.033), and/or DRD3 (P = 0.025) staining correlated negatively with the postoperative survival rate and positively with mortality. In competing-risks regression analysis, differentiation (P = 0.005), metastasis (P = 0.014), invasion (P = 0.028), AJCC stage (P = 0.003), Enneking stage (P = 0.036), and DAXX (P = 0.039), and DRD3(P = 0.019) expression were independent predictors of death from chondrosarcoma. The areas under receiver operating characteristic curves for DAXX, DRD3, and DISC1 expression were 0.673 (95% CI, 0.557–0.788; P = 0.010), 0.670 (95% CI, 0.556–0.784; P = 0.011), and 0.688 (95% CI, 0.573–0.802; P = 0.005), respectively. These results suggest that DAXX, DRD3, and DISC1 could serve as biomarkers of chondrosarcoma progression and predictors of its prognosis.

The Death-domain associated protein 6 (DAXX) is an evolutionarily conserved and ubiquitously expressed multifunctional protein that is implicated in many cellular processes, including transcription, cellular proliferation, cell cycle regulation, Fas-induced apoptosis, and many other events. In the nucleus, DAXX interacts with transcription factors, epigenetic modifiers, and chromatin-remodeling proteins such as the transcription regulator ATRX—the α-thalassemia/mental retardation syndrome X-linked ATP-dependent helicase II. Accordingly, DAXX is considered one of the main players involved in chromatin silencing and one of the most important factors that maintain integrity of the genome. In this brief review, we summarize available data regarding the general and specific functions of DAXX in mammalian early development, with special emphasis on the function of DAXX as a chaperone of the histone variant H3.3. Since H3.3 plays a key role in the developmental processes, especially in the pronounced rearrangements of heterochromatin compartment during oogenesis and embryogenesis, DAXX can be considered as an important factor supporting proper development. Specifically, loss of DAXX affects the recruitment of ATRX, transcription of tandem repeats and telomere functions, which results in a decrease in the viability of early embryos.

Death domain-associated protein 6 (Daxx) is a multifunctional, ubiquitously expressed and highly conserved chaperone protein involved in numerous cellular processes, including apoptosis, transcriptional repression, and carcinogenesis. In 2015, we identified Daxx as an antiretroviral factor that interfered with HIV-1 replication by inhibiting the reverse transcription step. In the present study, we sought to unravel the molecular mechanism of Daxx-mediated restriction and, in particular, to identify the protein(s) that Daxx targets in order to achieve its antiviral activity. First, we show that the SUMO-interacting motif (SIM) located at the C-terminus of the protein is strictly required for Daxx to inhibit HIV-1 reverse transcription. By performing a quantitative proteomic screen combined with classical biochemical analyses, we found that Daxx associated with incoming HIV-1 cores through a SIM-dependent interaction with cyclophilin A (CypA) and capsid (CA). Daxx was found to reside within a multiprotein complex associated with viral capsids, also containing TNPO3, TRIM5α, and TRIM34. Given the well-known influence of these cellular factors on the stability of HIV-1 cores, we investigated the effect of Daxx on the cytoplasmic fate of incoming cores and found that Daxx prevented HIV-1 uncoating in a SIM-dependent manner. Altogether, our findings suggest that, by recruiting TNPO3, TRIM5α, and TRIM34 and possibly other proteins onto incoming HIV-1 cores through a SIM-dependent interaction with CA-bound CypA, Daxx increases their stability, thus preventing uncoating and reverse transcription. Our study uncovers a previously unknown function of Daxx in the early steps of HIV-1 infection and further illustrates how reverse transcription and uncoating are two tightly interdependent processes.

Chromatin remodeling is critical for the regulation of the DNA damage response. We highlight findings from our recent study showing that the deposition of the histone variant H3.3 by the alpha-thalassemia mental retardation X-linked protein (ATRX) and the death domain associated protein (DAXX) chromatin remodeling complex regulates DNA repair synthesis during homologous recombination.

The presence of an activating mutation of the Wnt/β-catenin signaling pathway is found in ~90% of colorectal cancer (CRC) cases. Death domain-associated protein (DAXX), a nuclear protein, interacts with β-catenin in CRC cells. We investigated DAXX expression in 106 matched sample pairs of CRC and adjacent normal tissue by Western blotting. This study evaluated DAXX expression and its clinical implications in CRC. The results revealed that DAXX expression was significantly lower in the patients with the positive serum carcinoembryonic antigen (CEA) screening results compared to the patients with negative CEA screening levels (p < 0.001). It has been reported that CD24 is a Wnt target in CRC cells. Here, we further revealed that DAXX expression was significantly correlated with CD24 expression (rho = 0.360, p < 0.001) in 106 patients. Consistent with this, in the CEA-positive subgroup, of which the carcinomas expressed DAXX at low levels, they were significantly correlated with CD24 expression (rho = 0.461, p < 0.005). Therefore, reduced DAXX expression is associated with reduced CD24 expression in CRC. Notably, in the Hct116 cells, DAXX knockdown using short-hairpin RNA against DAXX (shDAXX) not only caused significant cell proliferation, but also promoted metastasis. The DAXX-knockdown cells also demonstrated significantly decreased CD24 expression, however the intracellular localization of CD24 did not change. Thus, DAXX might be considered as a potential regulator of CD24 or β-catenin expression, which might be correlated with proliferative and metastatic potential of CRC.

Par-4 (prostate apoptosis response-4) protein was originally found upregulated in prostate tumor cells undergoing apoptosis. Then it was further identified as a proapoptotic protein upregulated both in normal and leukemic lymphocytes. The aim of our study was to assess PAR-4 protein expression in the B cells of CLL patients and to examine its relationship with the expression of other proteins involved in the apoptosis process, such as DAXX, ZIPK and BCL-2. We found a positive relationship between PAR-4 and BCL-2 protein expression. Additionally, there was a positive correlation between PAR-4 and both DAXX and ZIPK protein expression. The results of our research were also analyzed in association with the principal CLL prognostic factors. There was a positive correlation between the expression of PAR-4 protein and the lactate dehydrogenase (LDH) serum concentration (p < 0.005). The expression of PAR-4 protein in B cells correlated positively with the percentage of CD38+ cells (p < 0.05), as well as with CD38+/ZAP-70+ cells (p < 0.05). Moreover, we found a close relationship between LPL protein expression or LPL/ADAM29 MFI ratio and PAR-4 protein expression. Our results confirm the significance of apoptosis deregulation in CLL, and suggest a possible relationship between PAR-4 expression and the clinical course of the disease. This however requires further investigation. (Folia Histochemica et Cytobiologica 2011; Vol. 49, No. 1, pp. 98–103)

The promyelocytic leukaemia protein (PML) has been implicated in regulation of multiple key cellular functions, from transcription to calcium homeostasis. PML pleiotropic role is in part related to its ability to localise to both the nucleus and cytoplasm. In the nucleus, PML is known to regulate gene transcription, a role linked to its ability to associate with transcription factors as well as chromatin-remodellers. A new twist came from the discovery that the PML-interacting protein death-associated protein 6 (DAXX) acts as chaperone for the histone H3.3 variant. H3.3 is found enriched at active genes, centromeric heterochromatin and telomeres, and has been proposed to act as important carrier of epigenetic information. Our recent work has implicated DAXX in regulation of H3.3 loading and transcription in the central nervous system (CNS). Remarkably, driver mutations in H3.3 and/or its loading machinery have been identified in brain cancer, thus suggesting a role for altered H3.3 function/deposition in CNS tumourigenesis. Aberrant H3.3 deposition may also play a role in leukaemia pathogenesis, given DAXX role in PML-RARα-driven transformation and the identification of a DAXX missense mutation in acute myeloid leukaemia. This review aims to critically discuss the existing literature and propose new avenues for investigation.

Abstract Background DNA viruses, such as herpes simplex virus type 1 (HSV-1), Simian virus 40 (SV40), and Cytomegaloviruses (CMV), start their replicative processes and transcription at specific nuclear domains known as ND10 (nuclear domain 10, also called PML bodies). It has been previously determined that for HSV-1 and SV40, a short DNA sequence and its binding protein are required and sufficient for cell localization of viral DNA replication and gene transcription. Results Our recent observations provide evidence that a foreign (not endogenous) DNA/protein complex in the nucleus recruits ND10 proteins. First, the complexes formed from the bacterial lac operator DNA and its binding protein (lac repressor), or from HPV11 (human papillomavirus 11) origin DNA and its binding protein (E2), co-localized with different ND10 proteins. Second, the HSV-1 amplicon without inserted lac operator DNA repeats distributed in the nucleus randomly, whereas the amplicon with lac operator DNA repeats associated with ND10, suggesting that DNA-binding proteins are required to localize at ND10. The cellular intrinsic DNA/protein complex (as detected for U2 DNA) showed no association with ND10. Furthermore, our examination of PML−/−, Daxx−/−, and Sp100-negative cells led to our discovering that DNA/protein complexes recruit ND10 protein independently. Using the GFP-LacI/Operator system, we were able to direct the transfected DNA to ND10 and found that gene expression was significantly repressed when the transfected DNA was directed to ND10. Conclusion Taken together, the results suggest that cells recognize DNA/protein complexes through a mechanism that involves interaction with the ND10-associated proteins.