Experimental maps of DNA structure at nucleotide resolution distinguish intrinsic from protein-induced DNA deformations

Recognition of DNA by proteins depends on DNA sequence and structure. Often unanswered is whether the structure of naked DNA persists in a protein–DNA complex, or whether protein binding changes DNA shape. While X-ray structures of protein–DNA complexes are numerous, the structure of naked cognate DNA is seldom available experimentally. We present here an experimental and computational analysis pipeline that uses hydroxyl radical cleavage to map, at single-nucleotide resolution, DNA minor groove width, a recognition feature widely exploited by proteins. For 11 protein–DNA complexes, we compared experimental maps of naked DNA minor groove width with minor groove width measured from X-ray co-crystal structures. Seven sites had similar minor groove widths as naked DNA and when bound to protein. For four sites, part of the DNA in the complex had the same structure as naked DNA, and part changed structure upon protein binding. We compared the experimental map with minor groove patterns of DNA predicted by two computational approaches, DNAshape and ORChID2, and found good but not perfect concordance with both. This experimental approach will be useful in mapping structures of DNA sequences for which high-resolution structural data are unavailable. This approach allows probing of protein family-dependent readout mechanisms.National Institutes of Health [R01GM106056 to R.R., T.D.T.; U54CA121852 in part to T.D.T.]; Boston University Undergraduate Research Opportunities Program [Faculty Matching Grants to D.O. and Y.J.]; USC Graduate School [Research Enhancement Fellowship and Manning Endowed Fellowship to T.P.C.]. R.R. is an Alfred P. Sloan Research Fellow. Funding for open access charge: Boston University. (R01GM106056 - National Institutes of Health; U54CA121852 - National Institutes of Health; Boston University Undergraduate Research Opportunities Program; USC Graduate School; Boston University)https://academic.oup.com/nar/article/46/5/2636/4829691?searchresult=1https://academic.oup.com/nar/article/46/5/2636/4829691?searchresult=1Published versio

Generation of Genic Diversity among Streptococcus pneumoniae Strains via Horizontal Gene Transfer during a Chronic Polyclonal Pediatric Infection

Although there is tremendous interest in understanding the evolutionary roles of horizontal gene transfer (HGT) processes that occur during chronic polyclonal infections, to date there have been few studies that directly address this topic. We have characterized multiple HGT events that most likely occurred during polyclonal infection among nasopharyngeal strains of Streptococcus pneumoniae recovered from a child suffering from chronic upper respiratory and middle-ear infections. Whole genome sequencing and comparative genomics were performed on six isolates collected during symptomatic episodes over a period of seven months. From these comparisons we determined that five of the isolates were genetically highly similar and likely represented a dominant lineage. We analyzed all genic and allelic differences among all six isolates and found that all differences tended to occur within contiguous genomic blocks, suggestive of strain evolution by homologous recombination. From these analyses we identified three strains (two of which were recovered on two different occasions) that appear to have been derived sequentially, one from the next, each by multiple recombination events. We also identified a fourth strain that contains many of the genomic segments that differentiate the three highly related strains from one another, and have hypothesized that this fourth strain may have served as a donor multiple times in the evolution of the dominant strain line. The variations among the parent, daughter, and grand-daughter recombinant strains collectively cover greater than seven percent of the genome and are grouped into 23 chromosomal clusters. While capturing in vivo HGT, these data support the distributed genome hypothesis and suggest that a single competence event in pneumococci can result in the replacement of DNA at multiple non-adjacent loci

Academic Cancer Center Phase I Program Development

Multiple factors critical to the effectiveness of academic phase I cancer programs were assessed among 16 academic centers in the U.S. Successful cancer centers were defined as having broad phase I and I/II clinical trial portfolios, multiple investigator‐initiated studies, and correlative science. The most significant elements were institutional philanthropic support, experienced clinical research managers, robust institutional basic research, institutional administrative efforts to reduce bureaucratic regulatory delays, phase I navigators to inform patients and physicians of new studies, and a large cancer center patient base. New programs may benefit from a separate stand‐alone operation, but mature phase I programs work well when many of the activities are transferred to disease‐oriented teams. The metrics may be useful as a rubric for new and established academic phase I programs.This commentary assesses the factors necessary for the effectiveness of academic phase I cancer programs. The metrics presented here may be useful as a rubric for new and established programs.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/139928/1/onco12106-sup-0001-suppinfo1.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/139928/2/onco12106.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/139928/3/onco12106-sup-0002-suppinfo2.pd

On the Action of 5-Amino-Salicylic Acid and Sulfapyridine on M. avium including Subspecies paratuberculosis

BACKGROUND: Introduced in 1942, sulfasalazine (a conjugate of 5-aminosalicylic acid (5-ASA) and sulfapyridine) is the most prescribed medication used to treat "inflammatory" bowel disease (IBD.) Although controversial, there are increasingly compelling data that Mycobacterium avium subspecies paratuberculosis (MAP) may be an etiological agent in some or all of IBD. We have shown that two other agents used in the therapy of IBD (methotrexate and 6-MP) profoundly inhibit MAP growth. We concluded that their most plausible mechanism of action is as antiMAP antibiotics. We herein hypothesize that the mechanism of action of 5-ASA and/or sulfapyridine may also simply be to inhibit MAP growth. METHODOLOGY: The effect on MAP growth kinetics by sulfasalazine and its components were evaluated in bacterial culture of two strains each of MAP and M. avium, using a radiometric ((14)CO(2) BACTEC(R)) detection system that quantifies mycobacterial growth as arbitrary "growth index units" (GI). Efficacy data are presented as "percent decrease in cumulative GI" (%-DeltacGI). PRINCIPAL FINDINGS: There are disparate responses to 5-ASA and sulfapyridine in the two subspecies. Against MAP, 5-ASA is inhibitory in a dose-dependent manner (MAP ATCC 19698 46%-DeltacGI at 64 microg/ml), whereas sulfapyridine has virtually no effect. In contrast, against M. avium ATCC 25291, 5-ASA has no effect, whereas sulfapyridine (88%-DeltacGI at 4 microg/ml) is as effective as methotrexate, our positive control (88%-DeltacGI at 4 microg/ml). CONCLUSIONS: 5-ASA inhibits MAP growth in culture. We posit that, unknowingly, the medical profession has been treating MAP infections since sulfasalazine's introduction in 1942. These observations may explain, in part, why MAP has not previously been identified as a human pathogen. We conclude that henceforth in clinical trials evaluating antiMAP agents in IBD, if considered ethical, the use of 5-ASA (as well as methotrexate and 6-MP) should be excluded from control groups

Mismatch repair deficiency predicts response of solid tumors to PD-1 blockade.

The genomes of cancers deficient in mismatch repair contain exceptionally high numbers of somatic mutations. In a proof-of-concept study, we previously showed that colorectal cancers with mismatch repair deficiency were sensitive to immune checkpoint blockade with antibodies to programmed death receptor-1 (PD-1). We have now expanded this study to evaluate the efficacy of PD-1 blockade in patients with advanced mismatch repair-deficient cancers across 12 different tumor types. Objective radiographic responses were observed in 53% of patients, and complete responses were achieved in 21% of patients. Responses were durable, with median progression-free survival and overall survival still not reached. Functional analysis in a responding patient demonstrated rapid in vivo expansion of neoantigen-specific T cell clones that were reactive to mutant neopeptides found in the tumor. These data support the hypothesis that the large proportion of mutant neoantigens in mismatch repair-deficient cancers make them sensitive to immune checkpoint blockade, regardless of the cancers\u27 tissue of origin

An integrative multi-omics analysis to identify candidate DNA methylation biomarkers related to prostate cancer risk

Abstract: It remains elusive whether some of the associations identified in genome-wide association studies of prostate cancer (PrCa) may be due to regulatory effects of genetic variants on CpG sites, which may further influence expression of PrCa target genes. To search for CpG sites associated with PrCa risk, here we establish genetic models to predict methylation (N = 1,595) and conduct association analyses with PrCa risk (79,194 cases and 61,112 controls). We identify 759 CpG sites showing an association, including 15 located at novel loci. Among those 759 CpG sites, methylation of 42 is associated with expression of 28 adjacent genes. Among 22 genes, 18 show an association with PrCa risk. Overall, 25 CpG sites show consistent association directions for the methylation-gene expression-PrCa pathway. We identify DNA methylation biomarkers associated with PrCa, and our findings suggest that specific CpG sites may influence PrCa via regulating expression of candidate PrCa target genes

Germline variation at 8q24 and prostate cancer risk in men of European ancestry

Chromosome 8q24 is a susceptibility locus for multiple cancers, including prostate cancer. Here we combine genetic data across the 8q24 susceptibility region from 71,535 prostate cancer cases and 52,935 controls of European ancestry to define the overall contribution of germline variation at 8q24 to prostate cancer risk. We identify 12 independent risk signals for prostate cancer (p < 4.28 × 10−15), including three risk variants that have yet to be reported. From a polygenic risk score (PRS) model, derived to assess the cumulative effect of risk variants at 8q24, men in the top 1% of the PRS have a 4-fold (95%CI = 3.62–4.40) greater risk compared to the population average. These 12 variants account for ~25% of what can be currently explained of the familial risk of prostate cancer by known genetic risk factors. These findings highlight the overwhelming contribution of germline variation at 8q24 on prostate cancer risk which has implications for population risk stratification