Enhancer elements are a key regulatory feature of many important genes. Several general features including the presence of specific histone modifications are used to demarcate potentially active ...enhancers. Here we reveal that putative enhancers marked with H3 lysine 79 (H3K79) di or trimethylation (me2/3) (which we name H3K79me2/3 enhancer elements or KEEs) can be found in multiple cell types. Mixed lineage leukemia gene (MLL) rearrangements (MLL-r) such as MLL-AF4 are a major cause of incurable acute lymphoblastic leukemias (ALL). Using the DOT1L inhibitor EPZ-5676 in MLL-AF4 leukemia cells, we show that H3K79me2/3 is required for maintaining chromatin accessibility, histone acetylation and transcription factor binding specifically at KEEs but not non-KEE enhancers. We go on to show that H3K79me2/3 is essential for maintaining enhancer-promoter interactions at a subset of KEEs. Together, these data implicate H3K79me2/3 as having a functional role at a subset of active enhancers in MLL-AF4 leukemia cells.
Abstract Background A post‐operative pancreatic fistula (POPF) is a major cause of morbidity and mortality after a pancreaticoduodenectomy (PD). This systematic review aimed to identify all scoring ...systems to predict POPF after a PD, consider their clinical applicability and assess the study quality. Method An electronic search was performed of Medline (1946–2014) and EMBASE (1996–2014) databases. Results were screened according to Preferred Reporting Items for Systematic Reviews and Meta‐Analyses (PRISMA) guidelines, and quality assessed according to the QUIPS (quality in prognostic studies) tool. Results Six eligible scoring systems were identified. Five studies used the International Study Group on Pancreatic Fistula (ISGPF) definition. The proposed scores feature between two and five variables and of the 16 total variables, the majority (12) featured in only one score. Three scores could be fully completed pre‐operatively whereas 1 score included intra‐operative and two studies post‐operative variables. Four scores were internally validated and of these, two scores have been subject to subsequent multicentre review. The median QUIPS score was 38 out of 50 (range 16–50). Conclusion These scores show potential in calculating the individualized patient risk of POPF. There is, however, much variation in current scoring systems and further validation in large multicentre cohorts is now needed.
Abstract
Transcription enhancers are essential activators of V(D)J recombination that orchestrate non-coding transcription through complementary, unrearranged gene segments. How transcription is ...coordinately increased at spatially distinct promoters, however, remains poorly understood. Using the murine immunoglobulin lambda (Igλ) locus as model, we find that three enhancer-like elements in the 3′ Igλ domain, Eλ3–1, HSCλ1 and HSE-1, show strikingly similar transcription factor binding dynamics and close spatial proximity, suggesting that they form an active enhancer hub. Temporal analyses show coordinate recruitment of complementary V and J gene segments to this hub, with comparable transcription factor binding dynamics to that at enhancers. We find further that E2A, p300, Mediator and Integrator bind to enhancers as early events, whereas YY1 recruitment and eRNA synthesis occur later, corresponding to transcription activation. Remarkably, the interplay between sense and antisense enhancer RNA is central to both active enhancer hub formation and coordinate Igλ transcription: Antisense Eλ3–1 eRNA represses Igλ activation whereas temporal analyses demonstrate that accumulating levels of sense eRNA boost YY1 recruitment to stabilise enhancer hub/promoter interactions and lead to coordinate transcription activation. These studies therefore demonstrate for the first time a critical role for threshold levels of sense versus antisense eRNA in locus activation.
Graphical Abstract
Graphical Abstract
Aberrant enhancer activation is a key mechanism driving oncogene expression in many cancers. While much is known about the regulation of larger chromosome domains in eukaryotes, the details of ...enhancer-promoter interactions remain poorly understood. Recent work suggests co-activators like BRD4 and Mediator have little impact on enhancer-promoter interactions. In leukemias controlled by the MLL-AF4 fusion protein, we use the ultra-high resolution technique Micro-Capture-C (MCC) to show that MLL-AF4 binding promotes broad, high-density regions of enhancer-promoter interactions at a subset of key targets. These enhancers are enriched for transcription elongation factors like PAF1C and FACT, and the loss of these factors abolishes enhancer-promoter contact. This work not only provides an additional model for how MLL-AF4 is able to drive high levels of transcription at key genes in leukemia but also suggests a more general model linking enhancer-promoter crosstalk and transcription elongation.
Chromosome conformation capture (3C) methods measure the spatial proximity between DNA elements in the cell nucleus. Many methods have been developed to sample 3C material, including the Capture-C ...family of protocols. Capture-C methods use oligonucleotides to enrich for interactions of interest from sequencing-ready 3C libraries. This approach is modular and has been adapted and optimized to work for sampling of disperse DNA elements (NuTi Capture-C), including from low cell inputs (LI Capture-C), as well as to generate Hi-C like maps for specific regions of interest (Tiled-C) and to interrogate multiway interactions (Tri-C). We present the design, experimental protocol and analysis pipeline for NuTi Capture-C in addition to the variations for generation of LI Capture-C, Tiled-C and Tri-C data. The entire procedure can be performed in 3 weeks and requires standard molecular biology skills and equipment, access to a next-generation sequencing platform, and basic bioinformatic skills. Implemented with other sequencing technologies, these methods can be used to identify regulatory interactions and to compare the structural organization of the genome in different cell types and genetic models.
V(D)J recombination generates antigen receptor diversity by mixing and matching individual variable (V), diversity (D), and joining (J) gene segments. An obligate by-product of many of these ...reactions is the excised signal circle (ESC), generated by excision of the DNA from between the gene segments. Initially, the ESC was believed to be inert and formed to protect the genome from reactive broken DNA ends but more recent work suggests that the ESC poses a substantial threat to genome stability. Crucially, the recombinase re-binds to the ESC, which can result in it being re-integrated back into the genome, to cause potentially oncogenic insertion events. In addition, very recently, the ESC/recombinase complex was found to catalyze breaks at recombination signal sequences (RSSs) throughout the genome, via a "cut-and-run" mechanism. Remarkably, the ESC/recombinase complex triggers these breaks at key leukemia driver genes, implying that this reaction could be a significant cause of lymphocyte genome instability. Here, we explore these alternate pathways and discuss their relative dangers to lymphocyte genome stability.
V(D)J recombination is essential to generate antigen receptor diversity but is also a potent cause of genome instability. Many chromosome alterations that result from aberrant V(D)J recombination ...involve breaks at single recombination signal sequences (RSSs). A long-standing question, however, is how such breaks occur. Here, we show that the genomic DNA that is excised during recombination, the excised signal circle (ESC), forms a complex with the recombinase proteins to efficiently catalyze breaks at single RSSs both in vitro and in vivo. Following cutting, the RSS is released while the ESC-recombinase complex remains intact to potentially trigger breaks at further RSSs. Consistent with this, chromosome breaks at RSSs increase markedly in the presence of the ESC. Notably, these breaks co-localize with those found in acute lymphoblastic leukemia patients and occur at key cancer driver genes. We have named this reaction “cut-and-run” and suggest that it could be a significant cause of lymphocyte genome instability.
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•A complex between the recombination by-product and RAGs triggers multiple DNA breaks•The breaks co-localize with chromosome breakpoints in acute lymphoblastic leukemias•The breaks occur at many frequently mutated genes in acute lymphoblastic leukemia•Cut-and-run may underpin the most common types of lymphocyte chromosome instabilities
V(D)J recombination errors have long been associated with chromosome alterations in lymphoid cancers. Here, Kirkham et al. describe an unexpected reaction where the recombination by-product complexes with the RAG recombinase to trigger DNA breaks throughout the genome. Notably, these breaks co-localize with those found in acute lymphoblastic leukemia.
Despite making up more than half of new doctors, women are underrepresented in most surgical specialties. Various reasons have been suggested for this including issues with work-life balance, ...discrimination and a lack of female role models in the specialty. We sought to quantify the extent of gender discrimination in leadership roles in surgical societies in the UK.
All major Surgical Specialty Organisations were identified via the Royal College of Surgeons Website. Leadership and committee information was collected via organisation websites on 5th September 2018. All societies were then contacted requesting data including total membership, their stage of training and the gender split.
Of the twenty-four organisations contacted, eighteen were able to provide data. Women accounted for 11.8% (2446/20 803) of consultant and 34.3% (5267/15 366) of trainee members. 2/24 presidents; 3/26 of vice presidents; 18.1% (39/215) of executive committees and 13.5% (49/364) of wider committee members were female. Thirty-four committee members were not included as they were not surgeons (23 female; 11 male).
Despite accounting for 27% of the surgical workforce and indeed 24% of surgical society members, women account for only 2 of 24 Presidents and 18.1% (39/215) of the executive committees of surgical societies in the UK. Action should be taken so women are fairly represented in leadership roles in surgical societies with one of the benefits being more visible role models for prospective female surgeons.
•Females represent only 12% of consultant surgeons and 27% of the surgical workforce despite constituting 47% of UK doctors.•Female surgeons are significantly underrepresented in organisational leadership.•Females comprised 8.3% of presidents and 11.5% of vice-president roles.•18.1% of executive committees and 13.5% of wider committee members are women.•Females accounted for 11.8% of consultant and 34.3% of trainee members.
Cellular ontogeny and MLL breakpoint site influence the capacity of MLL-edited CD34+ hematopoietic cells to initiate and recapitulate infant patients' features in pro-B-cell acute lymphoblastic ...leukemia (B-ALL). We provide key insights into the leukemogenic determinants of MLL-AF4+ infant B-ALL.
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