Positional specificity of different transcription factor classes within enhancers

Gene expression is controlled by sequence-specific transcription factors (TFs), which bind to regulatory sequences in DNA. TF binding occurs in nucleosome-depleted regions of DNA (NDRs), which generally encompass regions with lengths similar to those protected by nucleosomes. However, less is known about where within these regions specific TFs tend to be found. Here, we characterize the positional bias of inferred binding sites for 103 TFs within ∼500,000 NDRs across 47 cell types. We find that distinct classes of TFs display different binding preferences: Some tend to have binding sites toward the edges, some toward the center, and some at other positions within the NDR. These patterns are highly consistent across cell types, suggesting that they may reflect TF-specific intrinsic structural or functional characteristics. In particular, TF classes with binding sites at NDR edges are enriched for those known to interact with histones and chromatin remodelers, whereas TFs with central enrichment interact with other TFs and cofactors such as p300. Our results suggest distinct regiospecific binding patterns and functions of TF classes within enhancers. Keywords: transcription factor binding; gene regulation; genomics; chromatin structureNational Human Genome Research Institute (U.S.) (Grant 2U54HG003067-10)National Institute of General Medical Sciences (U.S.) (Grant T32GM007753

Mining data from 1000 genomes to identify the causal variant in regions under positive selection

The human genome contains hundreds of regions in which the patterns of genetic variation indicate recent positive natural selection, yet for most of these the underlying gene and the advantageous mutation remain unknown. We recently reported the development of a method, Composite of Multiple Signals (CMS), that combines tests for multiple signals of natural selection and increases resolution by up to 100-fold

A Composite of Multiple Signals Distinguishes Causal Variants in Regions of Positive Selection

The human genome contains hundreds of regions whose patterns of genetic variation indicate recent positive natural selection, yet for most the underlying gene and the advantageous mutation remain unknown. We developed a method, composite of multiple signals (CMS), that combines tests for multiple signals of selection and increases resolution by up to 100-fold. By applying CMS to candidate regions from the International Haplotype Map, we localized population-specific selective signals to 55 kilobases (median), identifying known and novel causal variants. CMS can not just identify individual loci but implicates precise variants selected by evolution.Organismic and Evolutionary BiologyOther Research Uni

Identification and Functional Validation of the Novel Antimalarial Resistance Locus PF10_0355 in Plasmodium falciparum

The Plasmodium falciparum parasite's ability to adapt to environmental pressures, such as the human immune system and antimalarial drugs, makes malaria an enduring burden to public health. Understanding the genetic basis of these adaptations is critical to intervening successfully against malaria. To that end, we created a high-density genotyping array that assays over 17,000 single nucleotide polymorphisms (~1 SNP/kb), and applied it to 57 culture-adapted parasites from three continents. We characterized genome-wide genetic diversity within and between populations and identified numerous loci with signals of natural selection, suggesting their role in recent adaptation. In addition, we performed a genome-wide association study (GWAS), searching for loci correlated with resistance to thirteen antimalarials; we detected both known and novel resistance loci, including a new halofantrine resistance locus, PF10_0355. Through functional testing we demonstrated that PF10_0355 overexpression decreases sensitivity to halofantrine, mefloquine, and lumefantrine, but not to structurally unrelated antimalarials, and that increased gene copy number mediates resistance. Our GWAS and follow-on functional validation demonstrate the potential of genome-wide studies to elucidate functionally important loci in the malaria parasite genome.Bill & Melinda Gates FoundationEllison Medical FoundationExxon Mobil FoundationFogarty International CenterNational Institute of Allergy and Infectious Diseases (U.S.)Burroughs Wellcome FundDavid & Lucile Packard FoundationNational Science Foundation (U.S.). Graduate Research Fellowship Progra

Mapping Copy Number Variation by Population Scale Genome Sequencing

Genomic structural variants (SVs) are abundant in humans, differing from other forms of variation in extent, origin and functional impact. Despite progress in SV characterization, the nucleotide resolution architecture of most SVs remains unknown. We constructed a map of unbalanced SVs (that is, copy number variants) based on whole genome DNA sequencing data from 185 human genomes, integrating evidence from complementary SV discovery approaches with extensive experimental validations. Our map encompassed 22,025 deletions and 6,000 additional SVs, including insertions and tandem duplications. Most SVs (53%) were mapped to nucleotide resolution, which facilitated analysing their origin and functional impact. We examined numerous whole and partial gene deletions with a genotyping approach and observed a depletion of gene disruptions amongst high frequency deletions. Furthermore, we observed differences in the size spectra of SVs originating from distinct formation mechanisms, and constructed a map of SV hotspots formed by common mechanisms. Our analytical framework and SV map serves as a resource for sequencing-based association studies.Organismic and Evolutionary Biolog

Impact of COVID-19 on cardiovascular testing in the United States versus the rest of the world

Objectives: This study sought to quantify and compare the decline in volumes of cardiovascular procedures between the United States and non-US institutions during the early phase of the coronavirus disease-2019 (COVID-19) pandemic. Background: The COVID-19 pandemic has disrupted the care of many non-COVID-19 illnesses. Reductions in diagnostic cardiovascular testing around the world have led to concerns over the implications of reduced testing for cardiovascular disease (CVD) morbidity and mortality. Methods: Data were submitted to the INCAPS-COVID (International Atomic Energy Agency Non-Invasive Cardiology Protocols Study of COVID-19), a multinational registry comprising 909 institutions in 108 countries (including 155 facilities in 40 U.S. states), assessing the impact of the COVID-19 pandemic on volumes of diagnostic cardiovascular procedures. Data were obtained for April 2020 and compared with volumes of baseline procedures from March 2019. We compared laboratory characteristics, practices, and procedure volumes between U.S. and non-U.S. facilities and between U.S. geographic regions and identified factors associated with volume reduction in the United States. Results: Reductions in the volumes of procedures in the United States were similar to those in non-U.S. facilities (68% vs. 63%, respectively; p = 0.237), although U.S. facilities reported greater reductions in invasive coronary angiography (69% vs. 53%, respectively; p < 0.001). Significantly more U.S. facilities reported increased use of telehealth and patient screening measures than non-U.S. facilities, such as temperature checks, symptom screenings, and COVID-19 testing. Reductions in volumes of procedures differed between U.S. regions, with larger declines observed in the Northeast (76%) and Midwest (74%) than in the South (62%) and West (44%). Prevalence of COVID-19, staff redeployments, outpatient centers, and urban centers were associated with greater reductions in volume in U.S. facilities in a multivariable analysis. Conclusions: We observed marked reductions in U.S. cardiovascular testing in the early phase of the pandemic and significant variability between U.S. regions. The association between reductions of volumes and COVID-19 prevalence in the United States highlighted the need for proactive efforts to maintain access to cardiovascular testing in areas most affected by outbreaks of COVID-19 infection

Development of a glycoconjugate vaccine to prevent invasive Salmonella Typhimurium infections in sub-Saharan Africa

Invasive infections associated with non-typhoidal Salmonella (NTS) serovars Enteritidis (SE), Typhimurium (STm) and monophasic variant 1,4,[5],12:i:- are a major health problem in infants and young children in sub-Saharan Africa, and currently, there are no approved human NTS vaccines. NTS O-polysaccharides and flagellin proteins are protective antigens in animal models of invasive NTS infection. Conjugates of SE core and O-polysaccharide (COPS) chemically linked to SE flagellin have enhanced the anti-COPS immune response and protected mice against fatal challenge with a Malian SE blood isolate. We report herein the development of a STm glycoconjugate vaccine comprised of STm COPS conjugated to the homologous serovar phase 1 flagellin protein (FliC) with assessment of the role of COPS O-acetyls for functional immunity. Sun-type COPS conjugates linked through the polysaccharide reducing end to FliC were more immunogenic and protective in mice challenged with a Malian STm blood isolate than multipoint lattice conjugates (>95% vaccine efficacy [VE] versus 30-43% VE). Immunization with de-O-acetylated STm-COPS conjugated to CRM197 provided significant but reduced protection against STm challenge compared to mice immunized with native STm-COPS:CRM197 (63-74% VE versus 100% VE). Although OPS O-acetyls were highly immunogenic, post-vaccination sera that contained various O-acetyl epitope-specific antibody profiles displayed similar in vitro bactericidal activity when equivalent titers of anti-COPS IgG were assayed. In-silico molecular modeling further indicated that STm OPS forms a single dominant conformation, irrespective of O-acetylation, in which O-acetyls extend outward and are highly solvent exposed. These preclinical results establish important quality attributes for an STm vaccine that could be co-formulated with an SE-COPS:FliC glycoconjugate as a bivalent NTS vaccine for use in sub-Saharan Africa

Combinatorial gene regulation by transcription factors

Thesis: Ph. D., Massachusetts Institute of Technology, Department of Biology, 2019Cataloged from PDF of thesis. "The Table of Contents does not accurately represent the page numbering"--Disclaimer page.Includes bibliographical references.Combinatorial gene regulation is encoded in enhancers and promoters in the form of binding sites for transcription factors (TFs), which collaboratively recruit the transcriptional machinery and drive gene expression. Using high-throughput and quantitative technologies developed by our lab and others, we studied TF binding sites in enhancers from numerous different cell types and regulatory systems, shedding light general principles of motif composition and organization in typical cellular regulatory elements. We find extensive synergy between TF binding sites, some with organizational constraints and some with flexible positioning. We demonstrate that different TFs bind at distinct positions within regulatory elements, suggesting a new type of architectural constraint in enhancers. Importantly, our analysis of both TF organization and cooperativity revealed distinctive patterns that separates TFs into potential functional classes. Together, our results suggest a structure of the regulatory code at the level of TF function and generate new hypotheses about regiospecific binding patterns and functions of TF classes within enhancers.by Sharon R. Grossman.Ph. D.Ph.D. Massachusetts Institute of Technology, Department of Biolog