Highly connected 3D chromatin networks established by an oncogenic fusion protein shape tumor cell identity.

Cell fate transitions observed in embryonic development involve changes in three-dimensional genomic organization that provide proper lineage specification. Whether similar events occur within tumor cells and contribute to cancer evolution remains largely unexplored. We modeled this process in the pediatric cancer Ewing sarcoma and investigated high-resolution looping and large-scale nuclear conformation changes associated with the oncogenic fusion protein EWS-FLI1. We show that chromatin interactions in tumor cells are dominated by highly connected looping hubs centered on EWS-FLI1 binding sites, which directly control the activity of linked enhancers and promoters to establish oncogenic expression programs. Conversely, EWS-FLI1 depletion led to the disassembly of these looping networks and a widespread nuclear reorganization through the establishment of new looping patterns and large-scale compartment configuration matching those observed in mesenchymal stem cells, a candidate Ewing sarcoma progenitor. Our data demonstrate that major architectural features of nuclear organization in cancer cells can depend on single oncogenes and are readily reversed to reestablish latent differentiation programs

The chromatin landscape of primary synovial sarcoma organoids is linked to specific epigenetic mechanisms and dependencies.

Synovial sarcoma (SyS) is an aggressive mesenchymal malignancy invariably associated with the chromosomal translocation t(X:18; p11:q11), which results in the in-frame fusion of the BAF complex gene SS18 to one of three SSX genes. Fusion of SS18 to SSX generates an aberrant transcriptional regulator, which, in permissive cells, drives tumor development by initiating major chromatin remodeling events that disrupt the balance between BAF-mediated gene activation and polycomb-dependent repression. Here, we developed SyS organoids and performed genome-wide epigenomic profiling of these models and mesenchymal precursors to define SyS-specific chromatin remodeling mechanisms and dependencies. We show that SS18-SSX induces broad BAF domains at its binding sites, which oppose polycomb repressor complex (PRC) 2 activity, while facilitating recruitment of a non-canonical (nc)PRC1 variant. Along with the uncoupling of polycomb complexes, we observed H3K27me3 eviction, H2AK119ub deposition and the establishment of de novo active regulatory elements that drive SyS identity. These alterations are completely reversible upon SS18-SSX depletion and are associated with vulnerability to USP7 loss, a core member of ncPRC1.1. Using the power of primary tumor organoids, our work helps define the mechanisms of epigenetic dysregulation on which SyS cells are dependent

EWSR1-ATF1 dependent 3D connectivity regulates oncogenic and differentiation programs in Clear Cell Sarcoma.

Oncogenic fusion proteins generated by chromosomal translocations play major roles in cancer. Among them, fusions between EWSR1 and transcription factors generate oncogenes with powerful chromatin regulatory activities, capable of establishing complex gene expression programs in permissive precursor cells. Here we define the epigenetic and 3D connectivity landscape of Clear Cell Sarcoma, an aggressive cancer driven by the EWSR1-ATF1 fusion gene. We find that EWSR1-ATF1 displays a distinct DNA binding pattern that requires the EWSR1 domain and promotes ATF1 retargeting to new distal sites, leading to chromatin activation and the establishment of a 3D network that controls oncogenic and differentiation signatures observed in primary CCS tumors. Conversely, EWSR1-ATF1 depletion results in a marked reconfiguration of 3D connectivity, including the emergence of regulatory circuits that promote neural crest-related developmental programs. Taken together, our study elucidates the epigenetic mechanisms utilized by EWSR1-ATF1 to establish regulatory networks in CCS, and points to precursor cells in the neural crest lineage as candidate cells of origin for these tumors

Baseline factors associated with early and late death in intracerebral haemorrhage survivors

Background and purpose: The aim of this study was to determine whether early and late death are associated with different baseline factors in intracerebral haemorrhage (ICH) survivors. Methods: This was a secondary analysis of the multicentre prospective observational CROMIS‐2 ICH study. Death was defined as ‘early’ if occurring within 6 months of study entry and ‘late’ if occurring after this time point. Results: In our cohort (n = 1094), there were 306 deaths (per 100 patient‐years: absolute event rate, 11.7; 95% confidence intervals, 10.5–13.1); 156 were ‘early’ and 150 ‘late’. In multivariable analyses, early death was independently associated with age [per year increase; hazard ratio (HR), 1.05, P = 0.003], history of hypertension (HR, 1.89, P = 0.038), pre‐event modified Rankin scale score (per point increase; HR, 1.41, P < 0.0001), admission National Institutes of Health Stroke Scale score (per point increase; HR, 1.11, P < 0.0001) and haemorrhage volume >60 mL (HR, 4.08, P < 0.0001). Late death showed independent associations with age (per year increase; HR, 1.04, P = 0.003), pre‐event modified Rankin scale score (per point increase; HR, 1.42, P = 0.001), prior anticoagulant use (HR, 2.13, P = 0.028) and the presence of intraventricular extension (HR, 1.73, P = 0.033) in multivariable analyses. In further analyses where time was treated as continuous (rather than dichotomized), the HR of previous cerebral ischaemic events increased with time, whereas HRs for Glasgow Coma Scale score, National Institutes of Health Stroke Scale score and ICH volume decreased over time. Conclusions: We provide new evidence that not all baseline factors associated with early mortality after ICH are associated with mortality after 6 months and that the effects of baseline variables change over time. Our findings could help design better prognostic scores for later death after ICH

oriC Region and replication termination site, dif, of the Xanthomonas campestris pv. campestris 17 chromosome

A 13-kb DNA fragment containing oriC and the flanking genes thdF, orf900, yidC, rnpA, rpmH, oriC, dnaA, dnaN, recF, and gyrB was cloned from the gram-negative plant pathogen Xanthomonas campestris pv. campestris 17. These genes are conserved in order with other eubacterial oriC genes and code for proteins that share high degrees of identity with their homologues, except for orf900, which has a homologue only in Xylella fastidiosa. The dnaA/dnaN intergenic region (273 bp) identified to be the minimal oriC region responsible for autonomous replication has 10 pure AT clusters of four to seven bases and only three consensus DnaA boxes. These findings are in disagreement with the notion that typical oriCs contain four or more DnaA boxes located upstream of the dnaA gene. The X campestris pv. campestris 17 attB site required for site-specific integration of cloned fragments from filamentous phage phiLf replicative form DNA was identified to be a dif site on the basis of similarities in nucleotide sequence and function with the Escherichia coli dif site required for chromosome dimer resolution and whose deletion causes filamentation of the cells. The oriC and dif sites were located at 12:00 and 6:00, respectively, on the circular X. campestris pv. campestris 17 chromosome map, similar to the locations found for E. coli sites. Computer searches revealed the presence of both the dif site and XerC/XerD recombinase homologues in 16 of the 42 fully sequenced eubacterial genomes, but eight of the dif sites are located far away from the 6:00 point instead of being placed opposite the cognate oriC. The differences in the relative position suggest that mechanisms different from that of E. coli may participate in the control of chromosome replication

Induction of hepatotoxicity by sanguinarine is associated with oxidation of protein thiols and disturbance of mitochondrial respiration

Sanguinarine (SANG) has been suggested to be one of the principle constituents responsible for the toxicity of Argemone mexicana seed oil. In this study, we focused on the possible mechanism of SANG-induced hepatotoxicity. The serum asparatate aminotransferase (AST), alanine aminotransferase (ALT), and lactate dehydrogenase (LDH) activities, hepatic vacuolization, lipid accumulation and lipid peroxidation of the liver were increased, and triglyceride (TG) was decreased in SANG-treated mice (10 mg kg(-1) i.p.), indicating damage to the liver. SANG induced cell death and DNA fragmentation, in a concentration- (0-30 mu M) and time-dependent (0-24 h) manner, and the cytotoxicity of SANG (15 mu M) was accompanied by an increase in reactive oxygen species and a lessening in protein thiol content; these outcomes were reversed by glutathione, N-acetyl-L-cysteine and 1,4-dithiothretol, and slightly improved by other antioxidants in hepatocytes. SANG can affect the function of mitochondria, leading to the depletion of the mitochondrial membrane potential and adenosine 5'-triphosphate content of hepatocytes. SANG caused an upcoupling effect of the respiratory chain at lower concentrations, but inhibited the respiratory chain at higher concentrations in mitochondira isolated from rat liver. In conclusion, the data suggest that SANG is a liver toxin that induces cytotoxicity in liver cells, possibly through oxidation of protein thiols, resulting in oxidative stress on the cells and disturbance of mitochondrial function. Copyright (C) 2008 John Wiley & Sons, Ltd

Sequencing of E. coli strain UTI89 on multiple sequencing platforms

10.1186/s13104-020-05335-4BMC Research Notes13148

Chromosome map of Xanthomonas campestris pv. campestris 17 with locations of genes involved in xanthan gum synthesis and yellow pigmentation

No plasmid was detected in Xanthomonas campestris pv. campestris 17, a strain of the causative agent of black rot in cruciferous plants isolated in Taiwan. Its chromosome was cut by PacI, PmeI, and SwaI into five, two, and six fragments, respectively, and a size of 4.8 Mb was estimated by summing the fragment lengths in these digests. Based on the data obtained from partial digestion and Southern hybridization using probes common to pairs of the overlapping fragments or prepared from linking fragments, a circular physical map bearing the PacI, PmeI, and SwaI sites was constructed for the X campestris pv. campestris 17 chromosome. Locations of eight eps loci involved in exopolysaccharide (xanthan gum) synthesis, two rm operons each possessing an unique I-CeuI site, one pig cluster required for yellow pigmentation, and nine auxotrophic markers were determined, using mutants isolated by mutagenesis with Tn5(pfm)CmKm. This transposon contains a polylinker with sites for several rare-cutting restriction endonucleases located between the chloramphenicol resistance and kanamycin resistance (Km(r)) genes, which upon insertion introduced additional sites into the chromosome. The recA and tdh genes, with known sequences, were mapped by tagging with the polylinker-Km(r) segment from Tn5(pfm)CmKm. This is the first map for X. campestris and would be useful for genetic studies of this and related Xanthomonas species