Global reference mapping of human transcription factor footprints.

Combinatorial binding of transcription factors to regulatory DNA underpins gene regulation in all organisms. Genetic variation in regulatory regions has been connected with diseases and diverse phenotypic traits1, but it remains challenging to distinguish variants that affect regulatory function2. Genomic DNase I footprinting enables the quantitative, nucleotide-resolution delineation of sites of transcription factor occupancy within native chromatin3-6. However, only a small fraction of such sites have been precisely resolved on the human genome sequence6. Here, to enable comprehensive mapping of transcription factor footprints, we produced high-density DNase I cleavage maps from 243 human cell and tissue types and states and integrated these data to delineate about 4.5 million compact genomic elements that encode transcription factor occupancy at nucleotide resolution. We map the fine-scale structure within about 1.6 million DNase I-hypersensitive sites and show that the overwhelming majority are populated by well-spaced sites of single transcription factor-DNA interaction. Cell-context-dependent cis-regulation is chiefly executed by wholesale modulation of accessibility at regulatory DNA rather than by differential transcription factor occupancy within accessible elements. We also show that the enrichment of genetic variants associated with diseases or phenotypic traits in regulatory regions1,7 is almost entirely attributable to variants within footprints, and that functional variants that affect transcription factor occupancy are nearly evenly partitioned between loss- and gain-of-function alleles. Unexpectedly, we find increased density of human genetic variation within transcription factor footprints, revealing an unappreciated driver of cis-regulatory evolution. Our results provide a framework for both global and nucleotide-precision analyses of gene regulatory mechanisms and functional genetic variation.

Comparing Robotic, Thoracoscopic, and Open Segmentectomy: A National Cancer Database Analysis.

INTRODUCTION: Minimally invasive approaches to lung resection have become widely acceptable and more recently, segmentectomy has demonstrated equivalent oncologic outcomes when compared to lobectomy for early-stage non-small cell lung cancer (NSCLC). However, studies comparing outcomes following segmentectomy by different surgical approaches are lacking. Our objective was to investigate the outcomes of patients undergoing robotic, video-assisted thoracoscopic surgery (VATS), or open segmentectomy for NSCLC using the National Cancer Database. METHODS: NSCLC patients with clinical stage I who underwent segmentectomy from 2010 to 2016 were identified. After propensity-score matching (1:4:1), multivariate logistic regression analyses were performed to determine predictors of 30-d readmissions, 90-d mortality, and overall survival. RESULTS: 22,792 patients met study inclusion. After matching, approaches included robotic (n = 2493; 17%), VATS (n = 9972; 66%), and open (n = 2493; 17%). An open approach was associated with higher 30-d readmissions (7% open versus 5.5% VATS versus 5.6% robot, P = 0.033) and 90-d mortality (4.4% open versus 2.2% VATS versus 2.5% robot, P < 0.001). A robotic approach was associated with improved 5-y survival (50% open versus 58% VATS versus 63% robot, P < 0.001). CONCLUSIONS: For patients with clinical stage I NSCLC undergoing segmentectomy, compared to the open approach, a VATS approach was associated with lower 30-d readmission and 90-d mortality. A robotic approach was associated with improved 5-y survival compared to open and VATS approaches when matched. Additional studies are necessary to determine if unrecognized covariates contribute to these differences.

eFORGE v2.0: updated analysis of cell type-specific signal in epigenomic data.

SUMMARY: The Illumina Infinium EPIC BeadChip is a new high-throughput array for DNA methylation analysis, extending the earlier 450k array by over 400 000 new sites. Previously, a method named eFORGE was developed to provide insights into cell type-specific and cell-composition effects for 450k data. Here, we present a significantly updated and improved version of eFORGE that can analyze both EPIC and 450k array data. New features include analysis of chromatin states, transcription factor motifs and DNase I footprints, providing tools for epigenome-wide association study interpretation and epigenome editing. AVAILABILITY AND IMPLEMENTATION: eFORGE v2.0 is implemented as a web tool available from https://eforge.altiusinstitute.org and https://eforge-tf.altiusinstitute.org/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Identifying Asian American lung cancer disparities: A novel analytic approach.

Objective: Asian Americans include heterogeneous subpopulations with unique burden as the only racial group with cancer as the leading cause of death. The purpose of the study was to identify differences in clinical stage and survival of patients with lung cancer between Asian Americans and its subgroups relative to other racial groups. Methods: Patients with lung cancer from 2016 National Cancer Database were divided into East Asian, Southeast Asian, South Asian subgroups based on geographic origins, and a composite Asian American group with White non-Hispanic, Black, and Hispanic comparison groups. Columnar z score analysis with adjusted residuals was employed and the terms underrepresented and overrepresented were utilized to describe significant statistical findings. Results: A total of 825,448 patients were analyzed. Asian Americans were underrepresented relative to White non-Hispanics in all clinical stages except IIIB and IV. In clinical stage IV, Asian Americans (51.0%), East Asians (47.2%), Southeast Asians (57.4%), and South Asians (52.2%) were overrepresented relative to White non-Hispanics (42.2%) and Southeast Asians were overrepresented relative to East Asians and South Asians. For survival across all stages, Asian Americans were overrepresented relative to White non-Hispanics and Blacks, but in clinical stage IV, Southeast Asians (17.9%) were underrepresented relative to East Asians (26.0%) and South Asians (26.6%). Conclusions: This is the first study to address lung cancer disparity in Asian American subgroups employing a novel analytical approach. Asian American subgroups demonstrated more advanced lung cancer diagnosis yet higher survival compared with White non-Hispanics, Blacks, and/or Hispanics with differences between subgroups. Interplay of complex factors may contribute to Asian American health disparities.

The Society of Thoracic Surgeons Expert Consensus on the Multidisciplinary Management and Resectability of Locally Advanced Non-Small Cell Lung Cancer.

BACKGROUND: The contemporary management and resectability of locally advanced lung cancer are undergoing significant changes as new data emerge regarding immunotherapy and targeted treatments. The objective of this document is to review the literature and present consensus among a group of multidisciplinary experts to guide the determination of resectability and management of locally advanced non-small cell lung cancer (NSCLC) in the context of contemporary evidence. METHODS: The Society of Thoracic Surgeon Workforce on Thoracic Surgery assembled a multidisciplinary expert panel comprised of thoracic surgeons and medical and radiation oncologists with established expertise in the management of lung cancer. A focused literature review was performed, and expert consensus statements were developed using a modified Delphi process to address three major themes: (1) Assessing Resectability and Multidisciplinary Management of Locally Advanced Lung Cancer, (2) Neoadjuvant (including peri-operative) therapy, and (3) Adjuvant therapy. RESULTS: A consensus was reached on 19 recommendations. These consensus statements reflect updated insights on resectability and multidisciplinary management of locally advanced lung cancer based on the latest literature and current clinical experience, mainly focusing on the appropriateness of surgical therapy and emerging data regarding neoadjuvant and adjuvant therapies. CONCLUSIONS: Despite the complex decision-making process in managing locally advanced lung cancer, this expert panel agreed on several key recommendations. This document provides guidance for thoracic surgeons and other medical professionals in the optimal management of locally advanced lung cancer based on the most updated evidence and literature.

A Review of Robotic Thoracic Surgery Adoption and Future Innovations.

The embracing of new technology and surgical innovation has been one of the hallmarks of cardiothoracic surgery. The adoption of video-assisted thoracoscopic surgery was the paradigm-changing perspective shift that has enabled an entire culture of minimally invasive cardiothoracic surgery. From this foundation robotically assisted thoracic surgery has developed into mainstream practice. The aim of this review is to recount the technological and academic milestones of the past as well as describe future technological innovations that will shape the future of thoracic robotic surgery.

A novel approach to quantifying surgical workflow in robotic-assisted lobectomy.

INTRODUCTION: Understanding surgical workflow is critical for optimizing efficiencies and outcomes; however, most research evaluating workflow is impacted by observer subjectivity, limiting its reproducibility, scalability, and actionability. To address this, we developed a novel approach to quantitatively describe workflow within robotic-assisted lobectomy (RL). We demonstrate the utility of this approach by analysing features of surgical workflow that correlate with procedure duration. METHODS: RL was deconstructed into 12 tasks by expert thoracic surgeons. Task start and stop times were annotated across videos of 10 upper RLs (5 right and 5 left). Markov Networks were used to estimate both the likelihood of transitioning from one task to another and each task-transition entropy (i.e. complexity). Associations between the frequency with which each task was revisited intraoperatively and procedure duration were assessed using Pearson's correlation coefficient. RESULTS: Entropy calculations identified fissure dissection and hilar node dissection as tasks with especially complex transitions, while mediastinal lymph node dissection and division of pulmonary veins were less complex. The number of transitions to three tasks significantly correlated with case duration (fissure dissection (R = 0.69, p = 0.01), dissect arteries (R = 0.59, p = 0.03), and divide arteries (R = 0.63, p = 0.03)). CONCLUSION: This pilot demonstrates the feasibility of objectively quantifying workflow between RL tasks and introduces entropy as a new metric of task-transition complexity. These innovative measures of surgical workflow enable detailed characterization of a given surgery and might indicate behaviour that impacts case progression. We discuss how these measures can serve as a foundation and be combined with relevant clinical information to better understand factors influencing surgical inefficiency.

Objective performance indicators of cardiothoracic residents are associated with vascular injury during robotic-assisted lobectomy on porcine models.

Surgical training relies on subjective feedback on resident technical performance by attending surgeons. A novel data recorder connected to a robotic-assisted surgical platform captures synchronized kinematic and video data during an operation to calculate quantitative, objective performance indicators (OPIs). The aim of this study was to determine if OPIs during initial task of a resident's robotic-assisted lobectomy (RL) correlated with bleeding during the procedure. Forty-six residents from the 2019 Thoracic Surgery Directors Association Resident Boot Camp completed RL on an ex vivo perfused porcine model while continuous video and kinematic data were recorded. For this pilot study, RL was segmented into 12 tasks and OPIs were calculated for the initial major task. Cases were reviewed for major bleeding events and OPIs of bleeding cases were compared to those who did not. Data from 42 residents were complete and included in the analysis. 10/42 residents (23.8%) encountered bleeding: 10/40 residents who started with superior pulmonary vein exposure and 0/2 residents who started with pulmonary artery exposure. Twenty OPIs for both hands were assessed during the initial task. Six OPIs related to instrument usage or smoothness of motion were significant for bleeding. Differences were statistically significant for both hands (p < 0.05). OPIs showing bimanual asymmetry indicated lower proficiency. This study demonstrates that kinematic and video analytics can establish a correlation between objective performance metrics and bleeding events in an ex vivo perfused lobectomy. Further study could assist in the development of focused exercises and simulation on objective domains to help improve overall performance and reducing complications during RL.

Comparison of Global Evaluative Assessment of Robotic Surgery with objective performance indicators for the assessment of skill during robotic-assisted thoracic surgery.

BACKGROUND: The Global Evaluative Assessment of Robotic Skills is a popular but ultimately subjective assessment tool in robotic-assisted surgery. An alternative approach is to record system or console events or calculate instrument kinematics to derive objective performance indicators. The aim of this study was to compare these 2 approaches and correlate the Global Evaluative Assessment of Robotic Skills with different types of objective performance indicators during robotic-assisted lobectomy. METHODS: Video, system event, and kinematic data were recorded from the robotic surgical system during left upper lobectomy on a standardized perfused and pulsatile ex vivo porcine heart-lung model. Videos were segmented into steps, and the superior vein dissection was graded independently by 2 blinded expert surgeons with Global Evaluative Assessment of Robotic Skills. Objective performance indicators representing categories for energy use, event data, movement, smoothness, time, and wrist articulation were calculated for the same task and compared to Global Evaluative Assessment of Robotic Skills scores. RESULTS: Video and data from 51 cases were analyzed (44 fellows, 7 attendings). Global Evaluative Assessment of Robotic Skills scores were significantly higher for attendings (P < .05), but there was a significant difference in raters' scores of 31.4% (defined as >20% difference in total score). The interclass correlation was 0.44 for 1 rater and 0.61 for 2 raters. Objective performance indicators correlated with Global Evaluative Assessment of Robotic Skills to varying degrees. The most highly correlated Global Evaluative Assessment of Robotic Skills domain was efficiency. Instrument movement and smoothness were highly correlated among objective performance indicator categories. Of individual objective performance indicators, right-hand median jerk, an objective performance indicator of change of acceleration, had the highest correlation coefficient (0.55). CONCLUSION: There was a relatively poor overall correlation between the Global Evaluative Assessment of Robotic Skills and objective performance indicators. However, both appear strongly correlated for certain metrics such as efficiency and smoothness. Objective performance indicators may be a potentially more quantitative and granular approach to assessing skill, given that they can be calculated mathematically and automatically without subjective interpretation.

The Scan, the Needle, or the Knife? National Trends in Diagnosing Stage I Lung Cancer.

OBJECTIVE: Indeterminate lung nodules have been increasingly discovered since the expansion of lung cancer screening programs. The diagnostic approach for suspicious nodules varies based on institutional resources and preferences. The aim of this study is to analyze factors associated with diagnostic modalities used for early-stage non-small cell lung cancer (NSCLC). METHODS: The National Cancer Database was queried for all patients with stage I NSCLC from 2004 to 2015. Four diagnostic modalities were identified, including clinical radiography alone (CRA), bronchial cytology (BC), procedural biopsy (PB), and surgical biopsy (SB). A multivariable multinomial logistic regression was used to assess associations of patient demographics, cancer characteristics, and facility characteristics with these modalities. RESULTS: Of 250,614 patients, 4,233 (1.7%) had CRA, 5,226 (2.1%) had BC, 147,621 (59.9%) had PB, and 93,534 (37.3%) had SB. Older patients were more likely to receive CRA (adjusted odds ratio [ORadj] = 5.3) and less likely to receive SB (ORadj = 0.73). Black patients were less likely to receive SB (ORadj = 0.83) and more likely to receive BC (ORadj = 1.31). Private insurance was associated with SB (ORadj = 1.11), whereas Medicaid was associated with BC (ORadj = 1.21). Patients more than 50 miles from the facility were more likely to undergo SB (ORadj = 1.25 vs PB; ORadj = 1.30 vs CRA; ORadj = 1.38 vs BC). Patients receiving SB had shorter days from diagnosis to treatment (23.0 vs 53.5 to 64.7 for other modalities, P < 0.001). CONCLUSIONS: Diagnostic SB to confirm early-stage NSCLC was associated with younger age, greater travel distance, and shorter time to treatment in comparison with other modalities. Black race and non-private insurance were less likely to be associated with SB.

Bacterial two-component systems as sensors for synthetic biology applications.

Two-component systems (TCSs) are a ubiquitous family of signal transduction pathways that enable bacteria to sense and respond to diverse physical, chemical, and biological stimuli outside and inside the cell. Synthetic biologists have begun to repurpose TCSs for applications in optogenetics, materials science, gut microbiome engineering, and soil nutrient biosensing, among others. New engineering methods including genetic refactoring, DNA-binding domain swapping, detection threshold tuning, and phosphorylation cross-talk insulation are being used to increase the reliability of TCS sensor performance and tailor TCS signaling properties to the requirements of specific applications. There is now potential to combine these methods with large-scale gene synthesis and laboratory screening to discover the inputs sensed by many uncharacterized TCSs and develop a large new family of genetically-encoded sensors that respond to an unrivaled breadth of stimuli.

Short and long-term outcomes of surgical intervention for empyema in the post-fibrinolytic era.

BACKGROUND: Open window thoracostomy (OWT) is indicated for patients with bronchopleural fistula (BPF) or trapped lung in the setting of empyema refractory to non-surgical interventions. We investigated the role of OWT in the era of minimally invasive surgeries, endobronchial valves and fibrinolytic therapy. METHODS: A retrospective chart review of all patients who underwent OWT at a single institution from 2010 to 2020 was performed. Indications for the procedure as well as operative details and morbidity and mortality were evaluated to determine patient outcomes for OWT. RESULTS: Eighteen patients were identified for the study. The most common indication for OWT was post-resectional BPF (n = 9). Prior to OWT, n = 11 patients failed other surgical or minimally invasive interventions. Patient comorbidities were quantified with the Charlson Comorbidity index (n = 11 score ≥ 5, 10-year survival ≤21%). Three (16.7%) patients died < 30 days post-operatively and 12 (66%) patients were deceased by the study's end (overall survival 24.0 ± 32.2 months). Mean number of ribs resected were 2.5 ± 1.2 (range 1-6) with one patient having 6 ribs removed. Patients were managed with negative pressure wound therapy (n = 9) or Kerlix packing (n = 9). Eleven patients (61.6%) underwent delayed closure (mean time from index surgery to closure 4.8 ± 6.7 months). CONCLUSIONS: Our study illustrates the significant comorbidities of patients undergoing OWT, the poor outcomes therein, and pitfalls associated with this procedure. We show that negative pressure wound therapy can be utilized as potential way to obliterate the pleural space and manage an open chest in the absence of an airleak; however, OWT procedures continue to be extremely morbid.

How We Do It: Implementing a Virtual, Multi-Institutional Collaborative Education Model for the COVID-19 Pandemic and Beyond.

OBJECTIVE: To describe the implementation of a virtual, multi-institutional educational collaboration involving over 50 general surgery residency programs during the COVID-19 pandemic that enabled enhanced learning for surgical residents despite social-distancing requirements. DESIGN: Description of Virginia Commonwealth University's virtual educational augmentation program and the development of a collaborative teaching network during the coronavirus pandemic. SETTING: This collaboration was initiated by Virginia Commonwealth University's Department of Surgery, Richmond, VA, and grew to include general surgery residency programs from across the nation. PARTICIPANTS: General surgery residents and faculty from Departments of General Surgery were recruited locally via direct emails and nationally via the Association of Program Directors' listserv and Twitter. In total, 52 institutions participated from every part of the country. RESULTS: A virtual, multi-institutional collaborative lecture series was initiated that grew to involve over 50 general surgery residency programs, allowing for daily didactics by experts in their fields during the initial surge of the COVID-19 pandemic, while maintaining social distancing and the provision of essential clinical care. CONCLUSION: A multi-institutional collaboration enabled continued didactic education during the coronavirus pandemic, vastly broadening the expertise, scope and variety available to residents, while decreasing burden on faculty. We believe this can serve as a framework for future multi-institutional collaborations that extend beyond the COVID-19 era.

Dynamic Control of Gene Expression with Riboregulated Switchable Feedback Promoters.

One major challenge in synthetic biology is the deleterious impacts of cellular stress caused by expression of heterologous pathways, sensors, and circuits. Feedback control and dynamic regulation are broadly proposed strategies to mitigate this cellular stress by optimizing gene expression levels temporally and in response to biological cues. While a variety of approaches for feedback implementation exist, they are often complex and cannot be easily manipulated. Here, we report a strategy that uses RNA transcriptional regulators to integrate additional layers of control over the output of natural and engineered feedback responsive circuits. Called riboregulated switchable feedback promoters (rSFPs), these gene expression cassettes can be modularly activated using multiple mechanisms, from manual induction to autonomous quorum sensing, allowing control over the timing, magnitude, and autonomy of expression. We develop rSFPs in Escherichia coli to regulate multiple feedback networks and apply them to control the output of two metabolic pathways. We envision that rSFPs will become a valuable tool for flexible and dynamic control of gene expression in metabolic engineering, biological therapeutic production, and many other applications.

Vulnerability of cholecystokinin-expressing GABAergic interneurons in the unilateral intrahippocampal kainate mouse model of temporal lobe epilepsy.

Temporal lobe epilepsy (TLE) is characterized by recurrent spontaneous seizures and behavioral comorbidities. Reduced hippocampal theta oscillations and hyperexcitability that contribute to cognitive deficits and spontaneous seizures are present beyond the sclerotic hippocampus in TLE. However, the mechanisms underlying compromised network oscillations and hyperexcitability observed in circuits remote from the sclerotic hippocampus are largely unknown. Cholecystokinin (CCK)-expressing basket cells (CCKBCs) critically participate in hippocampal theta rhythmogenesis, and regulate neuronal excitability. Thus, we examined whether CCKBCs were vulnerable in nonsclerotic regions of the ventral hippocampus remote from dorsal sclerotic hippocampus using the intrahippocampal kainate (IHK) mouse model of TLE, targeting unilateral dorsal hippocampus. We found a decrease in the number of CCK+ interneurons in ipsilateral ventral CA1 regions from epileptic mice compared to those from sham controls. We also found that the number of boutons from CCK+ interneurons was reduced in the stratum pyramidale, but not in other CA1 layers, of ipsilateral hippocampus in epileptic mice, suggesting that CCKBCs are vulnerable. Electrical recordings showed that synaptic connectivity and strength from surviving CCKBCs to CA1 pyramidal cells (PCs) were similar between epileptic mice and sham controls. In agreement with reduced CCKBC number in TLE, electrical recordings revealed a significant reduction in amplitude and frequency of IPSCs in CA1 PCs evoked by carbachol (commonly used to excite CCK+ interneurons) in ventral CA1 regions from epileptic mice versus sham controls. These findings suggest that loss of CCKBCs beyond the hippocampal lesion may contribute to hyperexcitability and compromised network oscillations in TLE.

Management of anastomotic leaks after esophagectomy and gastric pull-up.

Anastomotic leak is one of the most feared complications of esophagectomy, leading to prolonged hospital stay, increased postoperative mortality, and additional cost both to the patient and the hospital. Historically, anastomotic leaks have been treated with several techniques including conservative measures, percutaneous or operative drainage, primary surgical repair with buttressing, T-tube drainage, or excision of the esophageal replacement conduit with end esophagostomy. With advances in treatment modalities, including endoscopic stenting, clips and suturing, endoluminal vacuum-assisted closure (EVAC), such leaks increasingly are being managed without operative re-intervention and with salvage of the esophageal replacement conduit. For the purposes of this review, we identified studies analyzing the management of postoperative leak after esophagectomy. We then compared the efficacy of the various newer modalities for closure of anastomotic leaks and gastric conduit defects. We found both esophageal stent and EVAC sponges are effective treatments for closure of anastomotic leak. The chosen treatment modality for salvage of the esophageal replacement conduit is entirely dependent on the patient's clinical status and the surgeon's preference and experience. Emerging endoscopic and endoluminal therapies have increased the armamentarium of tools the esophageal surgeon has to facilitate successful resolution of anastomotic leaks following esophagectomy with reconstruction. While some literature suggests that EVACs have a slightly superior result in conduit success, we question this endorsement as EVACs mostly are utilized for contained leaks, many of which may have healed with conservative measures. This poses a challenge as there is clearly a bias given patient selection.

Optogenetic control of gut bacterial metabolism to promote longevity.

Gut microbial metabolism is associated with host longevity. However, because it requires direct manipulation of microbial metabolism in situ, establishing a causal link between these two processes remains challenging. We demonstrate an optogenetic method to control gene expression and metabolite production from bacteria residing in the host gut. We genetically engineer an Escherichia coli strain that secretes colanic acid (CA) under the quantitative control of light. Using this optogenetically-controlled strain to induce CA production directly in the Caenorhabditis elegans gut, we reveal the local effect of CA in protecting intestinal mitochondria from stress-induced hyper-fragmentation. We also demonstrate that the lifespan-extending effect of this strain is positively correlated with the intensity of green light, indicating a dose-dependent CA benefit on the host. Thus, optogenetics can be used to achieve quantitative and temporal control of gut bacterial metabolism in order to reveal its local and systemic effects on host health and aging.

Utility of erector spinae plane block in thoracic surgery.

BACKGROUND: Thoracic surgeons have been incorporating enhanced recovery after surgery (ERAS) protocols into their practices, not only to reduce narcotic usage but also to improve complication rates and decrease lengths of stay. Here, we describe the utility of a regional block technique that can be used for patients undergoing urgent or elective thoracic surgical procedures or suffering from rib fractures. METHODS: We report our initial one-year experience with these erector spinae plane (ESP) blocks. RESULTS: ESP blocks were placed in 42 patients. The procedure was performed by a trained team of anesthesiologists and certified nurse practitioners. It included placement of a catheter on the ipsilateral chest, followed by a 20 ml of 0.2% ropivacaine bolus and continuous infusion. Patients were then followed by the regional team, as long as the catheter was in place. While it had some technical challenges, the block was effective in 83.3% of patients with no reported mortality or major complications. However, given the confounding factors of the study (such as simultaneous implementation of ERAS protocol) and heterogeneity of the patient population, a control group was difficult to ascertain and meaningful opioid consumption analysis was difficult to perform. CONCLUSIONS: Regional blocks, such as the ESP block, complement fundamental ERAS principles and serve as an adjunct to the available armamentarium for non-narcotic ways to control pain in thoracic surgical and chest trauma patients. Continued collaboration between the thoracic surgeons and anesthesiologists is needed for its success.

Global Regulatory DNA Potentiation by SMARCA4 Propagates to Selective Gene Expression Programs via Domain-Level Remodeling.

The human genome encodes millions of regulatory elements, of which only a small fraction are active within a given cell type. Little is known about the global impact of chromatin remodelers on regulatory DNA landscapes and how this translates to gene expression. We use precision genome engineering to reawaken homozygously inactivated SMARCA4, a central ATPase of the human SWI/SNF chromatin remodeling complex, in lung adenocarcinoma cells. Here, we combine DNase I hypersensitivity, histone modification, and transcriptional profiling to show that SMARCA4 dramatically increases both the number and magnitude of accessible chromatin sites genome-wide, chiefly by unmasking sites of low regulatory factor occupancy. By contrast, transcriptional changes are concentrated within well-demarcated remodeling domains wherein expression of specific genes is gated by both distal element activation and promoter chromatin configuration. Our results provide a perspective on how global chromatin remodeling activity is translated to gene expression via regulatory DNA.

The critical role of persistent sodium current in hippocampal gamma oscillations.

Gamma network oscillations in the brain are fast rhythmic network oscillations in the gamma frequency range (~30-100 Hz), playing key roles in the hippocampus for learning, memory, and spatial processing. There is evidence indicating that GABAergic interneurons, including parvalbumin-expressing basket cells (PVBCs), contribute to cortical gamma oscillations through synaptic interactions with excitatory cells. However, the molecular, cellular, and circuit underpinnings underlying generation and maintenance of cortical gamma oscillations are largely elusive. Recent studies demonstrated that intrinsic and synaptic properties of GABAergic interneurons and excitatory cells are regulated by a slowly inactivating or non-inactivating sodium current (i.e., persistent sodium current, INaP), suggesting that INaP is involved in gamma oscillations. Here, we tested whether INaP plays a role in hippocampal gamma oscillations using pharmacological, optogenetic, and electrophysiological approaches. We found that INaP blockers, phenytoin (40 µM and 100 µM) and riluzole (10 µM), reduced gamma oscillations induced by optogenetic stimulation of CaMKII-expressing cells in CA1 networks. Whole-cell patch-clamp recordings further demonstrated that phenytoin (100 µM) reduced INaP and firing frequencies in both PVBCs and pyramidal cells without altering threshold and amplitude of action potentials, but increased rheobase in both cell types. These results suggest that INaP in pyramidal cells and PVBCs is required for hippocampal gamma oscillations, supporting a pyramidal-interneuron network gamma model. Phenytoin-mediated modulation of hippocampal gamma oscillations may be a mechanism underlying its anticonvulsant efficacy, as well as its contribution to cognitive impairments in epilepsy patients.