DNA-Protein Interactions in High Definition

An elegant, genome-wide approach to define the precise DNA sequences bound by transcription factors has been developed by Rhee and Pugh

Locus-specific editing of histone modifications at endogenous enhancers using programmable TALE-LSD1 fusions

Mammalian gene regulation is dependent on tissue-specific enhancers that can act across large distances to influence transcriptional activity1-3. Mapping experiments have identified hundreds of thousands of putative enhancers whose functionality is supported by cell type–specific chromatin signatures and striking enrichments for disease-associated sequence variants4-11. However, these studies did not address the in vivo functions of the putative elements or their chromatin states and could not determine which genes, if any, a given enhancer regulates. Here we present a strategy to investigate endogenous regulatory elements by selectively altering their chromatin state using programmable reagents. Transcription activator–like (TAL) effector repeat domains fused to the LSD1 histone demethylase efficiently remove enhancer-associated chromatin modifications from target loci, without affecting control regions. We find that inactivation of enhancer chromatin by these fusion proteins frequently causes down-regulation of proximal genes, revealing enhancer target genes. Our study demonstrates the potential of ‘epigenome editing’ tools to characterize an important class of functional genomic elements

Evidence-based Review on the Use of Proton Therapy in Lymphoma From the Particle Therapy Cooperative Group (PTCOG) Lymphoma Subcommittee.

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Strategic Operational Redesign Improves Prior Authorization Access: A Validation Study

Purpose: Obtaining prior authorization (PA) before treatment is becoming increasingly burdensome in oncology, especially in radiation oncology. Here, we describe the impact of a strategic novel operational PA redesign to shorten authorization time and to improve patient access to cancer care at a large United States academic proton therapy center. We ask whether such a redesign may be replicable and adoptable across oncology centers. Materials and Methods: Our PA redesign strategy was based on a 3-tiered approach. Specifically, we (1) held payors accountable to legally backed timelines, (2) leveraged expertise on insurance policies and practices, and (3) updated the submission, appeal writing, and planning procedures for PA. Metrics were compared at the following 3 time points: 6 months before, at phase-in, and at 6 months after intervention. Results: In analyzing the impact of improving PA access to care, the percentage of approvals for commercial proton beam therapy improved by an absolute 30.6% postintervention (P < .001). The proportion of commercially insured patients treated with proton beam therapy also increased by 6.2%, and the number of new starts rose by 11.7 patients/mo. Overall patient census increased by 13 patients/d. Median authorization time was 1 week, and 90% of surveyed providers reported reduced PA burden and improved patient care. Conclusion: This is the first validated, comprehensive operational strategy to improve access to cancer therapy while reducing the burden of PA. This novel approach may be helpful for addressing barriers to PA in medical and surgical oncology because the redesign is predicated on laws that regulate PA across disciplines

Functional Analysis of Human Hematopoietic Stem Cell Gene Expression Using Zebrafish

Although several reports have characterized the hematopoietic stem cell (HSC) transcriptome, the roles of HSC-specific genes in hematopoiesis remain elusive. To identify candidate regulators of HSC fate decisions, we compared the transcriptome of human umbilical cord blood and bone marrow CD34(+)CD33(−)CD38(−)Rho(lo)c-kit(+) cells, enriched for hematopoietic stem/progenitor cells with CD34(+)CD33(−)CD38(−)Rho(hi) cells, enriched in committed progenitors. We identified 277 differentially expressed transcripts conserved in these ontogenically distinct cell sources. We next performed a morpholino antisense oligonucleotide (MO)-based functional screen in zebrafish to determine the hematopoietic function of 61 genes that had no previously known function in HSC biology and for which a likely zebrafish ortholog could be identified. MO knock down of 14/61 (23%) of the differentially expressed transcripts resulted in hematopoietic defects in developing zebrafish embryos, as demonstrated by altered levels of circulating blood cells at 30 and 48 h postfertilization and subsequently confirmed by quantitative RT-PCR for erythroid-specific hbae1 and myeloid-specific lcp1 transcripts. Recapitulating the knockdown phenotype using a second MO of independent sequence, absence of the phenotype using a mismatched MO sequence, and rescue of the phenotype by cDNA-based overexpression of the targeted transcript for zebrafish spry4 confirmed the specificity of MO targeting in this system. Further characterization of the spry4-deficient zebrafish embryos demonstrated that hematopoietic defects were not due to more widespread defects in the mesodermal development, and therefore represented primary defects in HSC specification, proliferation, and/or differentiation. Overall, this high-throughput screen for the functional validation of differentially expressed genes using a zebrafish model of hematopoiesis represents a major step toward obtaining meaningful information from global gene profiling of HSCs

Ser ou não ser Mãe/Pai? Eis a questão –Motivações para a parentalidade

Background Local and/or regional recurrence and metachronous primary tumor arising in a previously irradiated area are rather frequent events in patients with head and neck squamous cell carcinoma (HNSCC). Re‐treatment is associated with an increased risk of serious toxicity and impaired quality of life (QOL) with an uncertain survival advantage. Methods We analyzed the literature on the efficacy and toxicity of photon/electron‐based external beam reirradiation for previously irradiated patients with HNSCC of non‐nasopharyngeal origin. Studies were grouped according to the radiotherapy technique used for reirradiation. Patient selection criteria, target volume identification method, tumor dose, fractionation schedule, systemic therapy administration, and toxicities were reviewed. Results In addition to disease‐related factors, current comorbidities and preexisting organ dysfunction must be considered when selecting patients for reirradiation. As morbidity from re‐treatment may be considerable and differ depending on which mode of re‐treatment is used, it is important to give patients information on potential morbidity outcomes so that an informed choice can be made within a shared decision‐making context. With improved dose distribution and adequate imaging support, including positron emission tomography‐CT, modern radiotherapy techniques may improve local control and reduce toxicity of reirradiation. A reirradiation dose of ≥60 Gy and a volume encompassing the gross tumor with up to a 5‐mm margin are recommended. Concomitant administration of systemic therapeutics and reirradiation is likely to be of similar benefit as observed in large randomized studies of upfront therapy. Conclusion Reirradiation, administered either with or without concurrent systemic therapy, is feasible and tolerable in properly selected patients with recurrent or a new primary tumor in a previously irradiated area of the head and neck, offering a meaningful survival (in the range of 10% to 30% at 2 years). Whenever feasible, salvage surgery is the method of choice for curative intent; patients at high‐risk for local recurrence should be advised that postoperative reirradiation is expected to increase locoregional control at the expense of higher toxicity and without survival advantage compared to salvage surgery without reirradiation. © 2014 Wiley Periodicals, Inc. Head Neck 37 : 134–150, 2015Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/110100/1/hed23542.pd

Merging paleobiology with conservation biology to guide the future of terrestrial ecosystems

Conservation of species and ecosystems is increasingly difficult because anthropogenic impacts are pervasive and accelerating. Under this rapid global change, maximizing conservation success requires a paradigm shift from maintaining ecosystems in idealized past states toward facilitating their adaptive and functional capacities, even as species ebb and flow individually. Developing effective strategies under this new paradigm will require deeper understanding of the long-term dynamics that govern ecosystem persistence and reconciliation of conflicts among approaches to conserving historical versus novel ecosystems. Integrating emerging information from conservation biology, paleobiology, and the Earth sciences is an important step forward on the path to success. Maintaining nature in all its aspects will also entail immediately addressing the overarching threats of growing human population, overconsumption, pollution, and climate change.Peer reviewe

Occupancy maps of 208 chromatin-associated proteins in one human cell type

Transcription factors are DNA-binding proteins that have key roles in gene regulation. Genome-wide occupancy maps of transcriptional regulators are important for understanding gene regulation and its effects on diverse biological processes. However, only a minority of the more than 1,600 transcription factors encoded in the human genome has been assayed. Here we present, as part of the ENCODE (Encyclopedia of DNA Elements) project, data and analyses from chromatin immunoprecipitation followed by high-throughput sequencing (ChIP–seq) experiments using the human HepG2 cell line for 208 chromatin-associated proteins (CAPs). These comprise 171 transcription factors and 37 transcriptional cofactors and chromatin regulator proteins, and represent nearly one-quarter of CAPs expressed in HepG2 cells. The binding profiles of these CAPs form major groups associated predominantly with promoters or enhancers, or with both. We confirm and expand the current catalogue of DNA sequence motifs for transcription factors, and describe motifs that correspond to other transcription factors that are co-enriched with the primary ChIP target. For example, FOX family motifs are enriched in ChIP–seq peaks of 37 other CAPs. We show that motif content and occupancy patterns can distinguish between promoters and enhancers. This catalogue reveals high-occupancy target regions at which many CAPs associate, although each contains motifs for only a minority of the numerous associated transcription factors. These analyses provide a more complete overview of the gene regulatory networks that define this cell type, and demonstrate the usefulness of the large-scale production efforts of the ENCODE Consortium

Locus-specific epigenetic remodeling controls addiction- and depression-related behaviors

Chronic exposure to drugs of abuse or stress regulates transcription factors, chromatin-modifying enzymes and histone post-translational modifications in discrete brain regions. Given the promiscuity of the enzymes involved, it has not yet been possible to obtain direct causal evidence to implicate the regulation of transcription and consequent behavioral plasticity by chromatin remodeling that occurs at a single gene. We investigated the mechanism linking chromatin dynamics to neurobiological phenomena by applying engineered transcription factors to selectively modify chromatin at a specific mouse gene in vivo. We found that histone methylation or acetylation at the Fosb locus in nucleus accumbens, a brain reward region, was sufficient to control drug- and stress-evoked transcriptional and behavioral responses via interactions with the endogenous transcriptional machinery. This approach allowed us to relate the epigenetic landscape at a given gene directly to regulation of its expression and to its subsequent effects on reward behavior