DNA N(6)-methyladenine (6mA) modification is commonly found in microbial genomes and plays important functions in regulating numerous biological processes in bacteria. However, whether 6mA occurs and what its potential roles are in higher-eukaryote cells remain unknown. Here, we show that 6mA is present in Drosophila genome and that the 6mA modification is dynamic and is regulated by the Drosophila Tet homolog, DNA 6mA demethylase (DMAD), during embryogenesis. Importantly, our biochemical assays demonstrate that DMAD directly catalyzes 6mA demethylation in vitro. Further genetic and sequencing analyses reveal that DMAD is essential for development and that DMAD removes 6mA primarily from transposon regions, which correlates with transposon suppression in Drosophila ovary. Collectively, we uncover a DNA modification in Drosophila and describe a potential role of the DMAD-6mA regulatory axis in controlling development in higher eukaryotes.

Metastatic disease is the primary cause of death in breast cancer, the most common malignancy in Western women. Loss of E-cadherin is associated with tumor metastasis, as well as with invasive lobular carcinoma (ILC), which accounts for 10%-15% of all breast cancers. To study the role of E-cadherin in breast oncogenesis, we have introduced conditional E-cadherin mutations into a mouse tumor model based on epithelium-specific knockout of p53. Combined loss of E-cadherin and p53 resulted in accelerated development of invasive and metastatic mammary carcinomas, which show strong resemblance to human ILC. Moreover, loss of E-cadherin induced anoikis resistance and facilitated angiogenesis, thus promoting metastatic disease. Our results suggest that loss of E-cadherin contributes to both mammary tumor initiation and metastasis.

Women carrying germ-line mutations in BRCA1 are strongly predisposed to developing breast cancers with characteristic features also observed in sporadic basal-like breast cancers. They appear as high-grade tumors with high proliferation rates and pushing borders. On the molecular level, they are negative for hormone receptors and ERBB2, display frequent TP53 mutations, and express basal epithelial markers. To study the role of BRCA1 and P53 loss of function in breast cancer development, we generated conditional mouse models with tissue-specific mutation of Brca1 and/or p53 in basal epithelial cells. Somatic loss of both BRCA1 and p53 resulted in the rapid and efficient formation of highly proliferative, poorly differentiated, estrogen receptor-negative mammary carcinomas with pushing borders and increased expression of basal epithelial markers, reminiscent of human basal-like breast cancer. BRCA1-and p53-deficient mouse mammary tumors exhibit dramatic genomic instability, and their molecular signatures resemble those of human BRCA1-mutated breast cancers. Thus, these tumors display important hallmarks of hereditary breast cancers in BRCA1-mutation carriers.mouse models ͉ conditional knockout G erm-line mutations in the human breast cancer susceptibility gene BRCA1 are responsible for 40% to 50% of hereditary breast cancers and confer increased risk for development of ovarian, colon, and prostate cancers (1, 2). BRCA1 has been implicated in various cellular processes, including maintenance of genome integrity, DNA replication and repair, chromatin remodeling, and transcriptional regulation (3, 4). Although the exact mechanism of mammary tumor suppression by BRCA1 remains largely unknown, cells with dysfunctional BRCA1 show defects in survival and proliferation, increased radiosensitivity, chromosomal abnormalities, G 2 /M checkpoint loss, and impaired homologous recombination repair (5).BRCA1-mutated breast cancers that arise in women with germline mutations in BRCA1 are high-grade, hormone receptornegative breast carcinomas with frequent mutation of TP53 (4, 6). They also possess a basal-like phenotype as defined by the expression of markers that are typical for basal/myoepithelial cells, such as the basal cytokeratins (CKs) CK5/6 and CK14 (7). Indeed, strong molecular similarities are observed between hereditary BRCA1-mutated breast cancers and sporadic basal-like breast carcinomas (8,9). This phenotypic overlap has led to the hypothesis that sporadic basal-like cancers may have defects in BRCA1-related pathways, such as the amplification of EMSY and the methylation of BRCA1 and FANCF (10).Despite the fact that several mouse strains with conventional or conditional mutations in Brca1 have been generated (11), no good mouse model for BRCA1-mutated basal-like breast cancer has been developed so far. Most conventional Brca1 knockouts are embryonic-lethal when bred to homozygosity, yet heterozygous ⌬11 allele, which encodes BRCA1-⌬11, a naturally occurring splice variant of Brca1 (19). Mouse mammary tumor models ba...

Background: Currently, the epidemic of coronavirus disease 2019 (COVID-19) has begun to spread worldwide. We aim to explore reliable evidence for the diagnosis and treatment of the COVID-19 by analyzing all the published studies by Chinese scholars on the clinical and imaging features in novel coronavirus pneumonia caused by SARS-CoV-2.Methods: We searched five medical databases including two Chinese and three English databases for all published articles on COVID-19 since the outbreak.A random-effects model was designed, and the imaging and clinical data from all studies were collected for meta-analysis.Results: Overall, 31 articles and 46 959 patients were included, including 10 English articles and 21 Chinese articles. The results of meta-analysis showed that the most common clinical manifestations were fever (87.3%; 0.838-0.909), cough (58.1%; 0.502-0.660), dyspnea (38.3%; 0.246-0.520), muscle soreness or fatigue (35.5%; 0.253-0.456), and chest distress (31.2%; −0.024 to 0.648). The main imaging findings were bilateral pneumonia (75.7%; 0.639-0.871) and ground-glass opacification (69.9%; 0.602-0.796). Among the patients, the incidence that required intensive care unit (ICU) was (29.3%; 0.190-0.395), the incidence with acute respiratory distress syndrome was (28.8%; 0.147-0.429), the incidence with multiple organ dysfunction syndrome was (8.5%; −0.008 to 0.179), and the case fatality rate of patients with COVID-19 was (6.8%; 0.044-0.093).

We have studied in vivo responses of ''spontaneous'' Brca1-and p53-deficient mammary tumors arising in conditional mouse mutants to treatment with doxorubicin, docetaxel, or cisplatin. Like human tumors, the response of individual mouse tumors varies, but eventually they all become resistant to the maximum tolerable dose of doxorubicin or docetaxel. The tumors also respond well to cisplatin but do not become resistant, even after multiple treatments in which tumors appear to regrow from a small fraction of surviving cells. Classical biochemical resistance mechanisms, such as up-regulated drug transporters, appear to be responsible for doxorubicin resistance, rather than alterations in drug-damage effector pathways. Our results underline the promise of these mouse tumors for the study of tumor-initiating cells and of drug therapy of human cancer.multidrug resistance ͉ P-glycoprotein ͉ cancer stem cells

Purpose: To investigate the clinical relevance of the recently characterized human oncoprotein cancerous inhibitor of protein phosphatase 2A (CIP2A) in human breast cancer. Experimental Design: CIP2A expression (mRNA and protein) was measured in three different sets of human mammary tumors and compared with clinicopathologic variables. The functional role of CIP2A in breast cancer cells was evaluated by small interfering RNA-mediated depletion of the protein followed by an analysis of cell proliferation, migration, anchorage-independent growth, and xenograft growth. Results: CIP2A mRNA is overexpressed (n = 159) and correlates with higher ScarffBloom-Richardson grades (n = 251) in samples from two independent human breast cancer patients. CIP2A protein was found to be overexpressed in 39% of 33 human breast cancer samples. Furthermore, CIP2A mRNA expression positively correlated with lymph node positivity of the patients and with the expression of proliferation markers and p53 mutations in the tumor samples. Moreover, CIP2A protein expression was induced in breast cancer mouse models presenting mammary gland-specific depletion of p53 and either BRCA1 or BRCA2. Functionally, CIP2A depletion was shown to inhibit the expression of its target protein c-Myc. Loss of CIP2A also inhibited anchorageindependent growth in breast cancer cells. Breast cancer is the most common malignancy that affects women, with >1 million cases occurring worldwide annually. Further, breast cancer is the most important cause of cancerrelated deaths in women. However, the understanding of the molecular mechanisms that maintain the malignant growth of breast cancer cells remains incomplete (1).The oncogenic transformation of human cells requires the perturbation of a distinct set of oncogenes and tumor suppressors (2). It was recently shown that the tumor suppressor activity of protein phosphatase 2A (PP2A) prevents the transformation of human breast epithelial cells (3). The role of PP2A as a relevant breast cancer tumor suppressor was further strengthened by a recent study showing that somatic mutations occurred in one of the subunits of the functional PP2A trimer (PP2A Aβ) in 13% of human breast cancers and that PP2A trimers containing this mutation fail to suppress the oncogenic activity of RalA (4, 5). In

Purpose: To assess efficacy of the novel, selective poly(ADP-ribose) polymerase-1 (PARP-1) inhibitor AZD2281against newly established BRCA2-deficient mouse mammary tumor cell lines and to determine potential synergy between AZD2281and cisplatin. Experimental Design: We established and thoroughly characterized a panel of clonal cell lines from independent BRCA2-deficient mouse mammary tumors and BRCA2-proficient control tumors. Subsequently, we assessed sensitivity of these lines to conventional cytotoxic drugs and the novel PARP inhibitor AZD2281. Finally, in vitro combination studies were done to investigate interaction between AZD2281and cisplatin. Results: Genetic, transcriptional, and functional analyses confirmed the successful isolation of BRCA2-deficient and BRCA2-proficient mouse mammary tumor cell lines.Treatment of these cell lines with 11 different anticancer drugs or with g-irradiation showed that AZD2281, a novel and specific PARP inhibitor, caused the strongest differential growth inhibition of BRCA2-deficient versus BRCA2-proficient mammary tumor cells. Finally, drug combination studies showed synergistic cytotoxicity of AZD2281 and cisplatin against BRCA2-deficient cells but not against BRCA2-proficient control cells. Conclusion: We have successfully established the first set of BRCA2-deficient mammary tumor cell lines, which form an important addition to the existing preclinical models for BRCA-mutated breast cancer. The exquisite sensitivity of these cells to the PARP inhibitor AZD2281, alone or in combination with cisplatin, provides strong support for AZD2281as a novel targeted therapeutic against BRCA-deficient cancers.

Interleukin-12 family cytokines have emerged as critical regulators of immunity with some members (IL-12, IL-23) associated with disease pathogenesis while others (IL-27sue injury by promoting the expansion of regulatory B and T-cell subsets [2,3].Discovery of IL-23 in 2000 [4] led to the reevaluation of IL-12 and IL-23 in autoimmune diseases. For example, therapeutic targeting of IL-12p40 decreases pathology in many mouse models of autoimmune diseases [5], while disease is exacerbated in IL-12p35-deficient mice [6,7]. Thus, IL-23 rather than IL-12 was * These authors contributed equally to this work as first authors.* * These authors contributed equally to this work as senior authors. Eur. J. Immunol. 2016. 46: 1343-1350 found to be the critical cytokine for autoimmune inflammation including experimental immune-mediated disease [6][7][8][9][10]. Currently, at least ten therapeutic agents targeting IL-23 are being tested in the clinic for more than 17 human immune-mediated diseases [11]. Both IL-27 and IL-35 have immune-suppressive activities and are also cytokines with strikingly diverse influences on the immune response so that viable therapeutic targets may also be exploited for treatment of human inflammatory diseases [12,13]. Thus, understanding immunobiology of IL-12 family cytokines would undoubtedly provide valuable knowledge that can be exploited therapeutically. The IL-12 family cytokines are α/β heterodimers consisting of one α subunit (IL-23p19, IL-27p28, IL-12p35) and one β chain (IL-12p40, Ebi3) [14,15]. Although there are currently four known members in the family, the predictable range of combinations is six and it is conceivable that additional pairings such as IL-23p19/Ebi3 are possible [12,[14][15][16][17]. In this study, we sought to discover additional IL-12 members that might exist in nature. By combining different alpha and beta IL-12 subunit proteins in vitro we detected a novel stable p19/Ebi3 heterodimeric complex by immunoprecipitation. We have characterized the p19/Ebi3 cytokine (IL-39) and demonstrated that it possesses biological activities in vitro and in vivo. Results IL-23p19 (p19) and Ebi3 form a composite factor (IL-39)To examine whether p19 can form a stable complex with Ebi3, we mixed equal amounts of the two proteins and immunoprecipitation (IP)/Western blot analyses revealed formation of a stable human p19/Ebi3 complex (Fig. 1A). We could not detect the p19/Ebi3 following IP with isotype IgG or anti-c-Jun antibody, providing suggestive evidence for potential bona fide p19/Ebi3 cytokine. To confirm our finding in another animal species, we genetically engineered and expressed mouse p19 and Ebi3 subunits in CHO cells (Fig. 1B). IP of supernatants derived from transfectants with anti-p19 mAb and followed by Western blot analysis using anti-Ebi3 mAb confirmed coexpression p19 and Ebi3 and formation of a stable p19/Ebi3 heterodimer (Fig. 1C). We further confirmed this observation by reciprocal IP with anti-Ebi3 mAb and Western blotting with anti-p19 mAb and the p19/Ebi3 comple...