The vaccinia virus (VV) E3L protein is essential for virulence and has anti-apoptotic activity. In mice, Z-DNA-binding activity of the N-terminal domain of E3L (Z␣) is necessary for viral lethality. Here, we report that inhibition of hygromycin-B-induced apoptosis in HeLa cells depends on Z-DNA binding of the E3L Z␣ domain. Z-DNA-binding domains of other proteins are equally effective in blocking apoptosis. Using a transient reporter assay, we demonstrate transactivation of human IL-6, nuclear factor of activated T cells (NF-AT), and p53 genes by E3L. This activation also requires Z-DNA binding of the N-terminal domain of E3L. Overall, this work suggests that the important role of E3L in VV pathogenesis involves modulating expression of host cellular genes at the transcriptional level and inhibiting apoptosis of host cells through Z-DNA binding.apoptosis ͉ interleukin-6 ͉ p53 ͉ poxvirus V accinia virus (VV) is a large double-stranded DNA virus encoding Ϸ190 genes, and it causes major changes in the host cell machinery shortly after infection. E3L is a host range gene, necessary for efficient VV replication in several cell lines (1) and is required for VV pathogenesis (2). The E3L gene is expressed early during infection, and the protein is present in both the nucleus and cytoplasm of infected and transfected cells. E3L accumulates in the nucleus (3), but little is known about its activities there. E3L has two domains, an N-terminal Z-DNAbinding protein domain (Z␣) and a C-terminal double-stranded RNA-binding domain (Fig. 1A). The N-terminal region is similar to the Z␣ domain of several Z-DNA-binding protein families that include the RNA editing enzyme ADAR1 (double-stranded RNA adenosine deaminase) (4), the tumor-related DLM1 (or ZBP1) protein (5), and the recently described PKR-like kinase of bony fish (6). The crystal structures of two Z␣ domains, Z␣ ADAR1 (Fig. 1B) (7) and Z␣ DLM1 (5), have shown them complexed to Z-DNA. Both N-terminal and C-terminal domains are required for infection and full pathogenesis in the mouse model (2). It has been shown that viral pathogenicity requires E3L binding to Z-DNA (8). The N-terminal half of the E3L protein is highly conserved among distantly related poxviruses, and E3L from another poxvirus has been cocrystallized with Z-DNA (9), but the functional role of this region has not been well characterized. It has been suggested that the N-terminal domain of E3L is involved in the direct inhibition of protein kinase R (PKR) activation, nuclear localization, and Z-DNA binding (8,10,11). A VV mutant lacking E3L induces apoptosis in HeLa cells (12). Furthermore, it has been reported that E3L inhibits dsRNA-induced apoptosis in NIH 3T3 cells and has some oncogenic properties (12). This suggests the possibility that E3L acts by modifying transcription, as do proteins from other DNA viruses (see Supporting Text, which is published as supporting information on the PNAS web site).In this report, we have inquired whether E3L functions as a transcriptional regulator on several genes tha...

In this article, the effect of a d(CG) DNA dinucleotide repeat sequence on RNA polymerase II transcription is examined in yeast Saccharomyces cerevisiae. Our previous report shows that a d(CG)n dinucleotide repeat sequence located proximally upstream of the TATA box enhances transcription from a minimal CYC1 promoter in a manner that depends on its surrounding negative supercoiling. Here, we demonstrate that the d(CG)9 repeat sequence stimulates gene activity by forming a Z-DNA secondary structure. Furthermore, the extent of transcriptional enhancement by Z-DNA is promoter-specific and determined by its separation distance relative to the TATA box. The stimulatory effect exerted by promoter proximal Z-DNA is not affected by helical phasing relative to the TATA box, suggesting that Z-DNA effects transcription without interacting with the general transcription machinery by loopingout the intervening DNA. A nucleosome-scanning assay reveals that the d(CG)9 repeat sequence in the Z conformation blocks nucleosome formation, and it is found in the linker DNA with two flanking nucleosomes. This result suggests that Z-DNA formation proximally upstream of a promoter is sufficient to demarcate the boundaries of its neighboring nucleosomes, which produces transcriptionally favorable locations for the TATA box near the nucleosomal DNA-entry site and at dyad positions on the nucleosome. These findings suggest that Z-DNA formation in chromatin is a part of the ''genomic code'' for nucleosome positioning in vivo.chromatin remodeling ͉ promoter ͉ transcription ͉ nucleosome positioning

Background Rivoceranib, a novel tyrosine kinase inhibitor, exhibits anti-tumour effects by selectively blocking vascular endothelial growth factor receptor-2 (VEGFR2) in cancer cells. Recently, the therapeutic effects of rivoceranib on solid tumours have been elucidated in human patients. However, the anti-tumour effects of rivoceranib against canine cancer remain unclear. Here, we investigated the anti-tumour effects of rivoceranib using in vitro and in vivo mouse xenograft models. Methods We performed cell proliferation, cell cycle, and migration assays to determine the effects of rivoceranib on canine solid tumour cell lines in vitro. Furthermore, apoptosis and angiogenesis in tumour tissues were examined using a TUNEL assay and immunohistochemistry methods with an anti-cluster of differentiation-31 antibody, respectively. Additionally, the expression levels of cyclin-D1 and VEGFR2 activity were determined using western blot analysis. Results Rivoceranib treatment showed anti-proliferative effects and mediated cell cycle arrest in the canine melanoma cell line (LMeC) and the mammary gland tumour (MGT) cell line (CHMp). In animal experiments, rivoceranib decreased the average volume of LMeC cells compared to that following control treatment, and similar results were observed in CHMp cells. Histologically, rivoceranib induced apoptosis and exerted an anti-angiogenic effect in tumour tissues. It also downregulated the expression of cyclin-D1 and inhibited VEGFR2 activity. Conclusion Our results show that rivoceranib inhibits proliferation and migration of tumour cells. These findings support the potential application of rivoceranib as a novel chemotherapeutic strategy for canine melanoma and MGTs.

Purpose: The Health Insurance Review and Assessment Service (HIRA) in South Korea initiated a quality assessment (QA) program for blood transfusion healthcare services in 2020 to ensure patient safety and appropriate blood use. This study examines the quality of blood transfusion services since the first national QA program for blood transfusion services in Korea.Methods: We analyzed HIRA claims and QA investigation data based on inpatient medical records from all tertiary, general, and primary hospitals between October 2020 and March 2021. The target population was patients aged 18 years and older who received either total knee arthroplasty or red blood cell transfusion. The QA indicators for transfusion healthcare service consisted of four quality indicators and four monitoring indicators.Results: We analyzed the results of QA indicators for transfusion service from the medical records of 189,668 patients from 1,171hospitals and expressed indicators as proportions. The average results for evaluation indicators were as follows: transfusion checklist presence, 64.8%; irregular antibody tests, 61.8%; transfusions in which the hemoglobin levels before transfusion met the transfusion guidelines for patients undergoing total knee arthroplasty, 20.6%, and transfusions in patients undergoing total knee arthroplasty, 59.3%. The average results for monitoring indicators were as follows: transfusion management implementation in medical institutions, 56.9%; preoperative anemia management in anemia patients undergoing total knee arthroplasty, 43.9%; one-unit transfusions, 82.5%; and the transfusion index.Conclusion: The quality of blood transfusion healthcare varied and the assessment revealed that there is scope for improvement. Hospitals require more effective blood transfusion management and this can be facilitated by providing feedback on the QA results about blood transfusion healthcare services to medical institutions, and by disclosing the results to the public.