Abstract
Human DNA ligase I (LIG1) is the main replicative ligase and it also seals DNA breaks to complete DNA repair and recombination pathways. Immune compromised patients harbor hypomorphic LIG1 ...alleles encoding substitutions of conserved arginine residues, R771W and R641L, that compromise LIG1 activity through poorly defined mechanisms. To understand the molecular basis of LIG1 syndrome mutations, we determined high resolution X-ray structures and performed systematic biochemical characterization of LIG1 mutants using steady-state and pre-steady state kinetic approaches. Our results unveil a cooperative network of plastic DNA-LIG1 interactions that connect DNA substrate engagement with productive binding of Mg2+ cofactors for catalysis. LIG1 syndrome mutations destabilize this network, compromising Mg2+ binding affinity, decreasing ligation efficiency, and leading to elevated abortive ligation that may underlie the disease pathology. These findings provide novel insights into the fundamental mechanism by which DNA ligases engage with a nicked DNA substrate, and they suggest that disease pathology of LIG1 syndrome could be modulated by Mg2+ levels.
DNA methylation variations are prevalent in human obesity but evidence of a causative role in disease pathogenesis is limited. Here, we combine epigenome-wide association and integrative genomics to ...investigate the impact of adipocyte DNA methylation variations in human obesity. We discover extensive DNA methylation changes that are robustly associated with obesity (N = 190 samples, 691 loci in subcutaneous and 173 loci in visceral adipocytes, P < 1 × 10-7). We connect obesity-associated methylation variations to transcriptomic changes at >500 target genes, and identify putative methylation-transcription factor interactions. Through Mendelian Randomisation, we infer causal effects of methylation on obesity and obesity-induced metabolic disturbances at 59 independent loci. Targeted methylation sequencing, CRISPR-activation and gene silencing in adipocytes, further identifies regional methylation variations, underlying regulatory elements and novel cellular metabolic effects. Our results indicate DNA methylation is an important determinant of human obesity and its metabolic complications, and reveal mechanisms through which altered methylation may impact adipocyte functions.
Clitellata is a major clade of Annelida comprising nearly all freshwater and terrestrial annelids as well as several marine species. We investigated clitellate phylogenetic relationships using ...transcriptomes sampled from 74 taxa (64 clitellates and 10 polychaetes), including multiple representatives of nearly all major clitellate higher taxa (Branchiobdellida, Capilloventridae, Crassiclitellata, Enchytraeidae, Haplotaxidae, Hirudinida, Lumbriculida, Moniligastridae, Naididae, Parvidrilidae, Phreodrilidae, Propappidae and Randiellidae). We used a number of filtered data matrices and phylogenetic analyses to examine the effects of data partitioning, missing data and compositional and branch‐length heterogeneity and used the resulting phylogenies for divergence time estimation and ancestral habitat reconstructions. All analyses and filtering methods produced a consistent, strongly supported topology in which (a) Enchytraeidae, Hirudinida, Hirudinea (here, Branchiobdellida plus Hirudinida), Lumbriculida, Lumbriculata (Lumbriculida plus Hirudinea), Phreodrilidae and Naididae are monophyletic, (b) a Parvidrilidae + Randiellidae clade is sister to the rest of Clitellata, (c) Phreodrilidae is sister to Naididae, (d) Haplotaxidae is non‐monophyletic, with some haplotaxids grouping with Crassiclitellata + Moniligastridae, (e) the Phreodrilidae + Naididae clade is sister to all other clitellates except Parvidrilidae + Randiellidae and Capilloventridae, and (f) Lumbriculata is sister to the Crassiclitellata + Moniligastridae + Haplotaxidae (in part) clade. Ancestral habitat reconstructions and divergence time analysis suggested that the most recent common ancestor of Clitellata lived in freshwater during the Devonian (419–359 million years ago) and that all major extant clitellate lineages arose over the next ~150 million years, with multiple lineages subsequently returning to marine habitats or invading land. This study provides a phylogenetic framework for further investigation of the geological, environmental and biotic forces and genomic changes that may have impacted clitellate evolution and enabled several major habitat transitions within this group.
Highlights • Individuals with HIV onantiretroviral treatment (ART) experience mortality at lower levels of alcohol use. • Individuals with HIV experience physiologic frailty at lower levels of ...alcohol use. • Alcohol consumption limits should be lower among HIV+ individuals.
What are the ethics of using voices generated by artificial intelligence or "deepfake" technology in documentary film? This case study explores the controversy surrounding the use of AI to ...reconstruct Anthony Bourdain's voice in the biographical film, Roadrunner.
Genetic screening of the breast and ovarian cancer susceptibility gene BRCA1 has uncovered a large number of variants of uncertain clinical significance. Here, we use biochemical and cell-based ...transcriptional assays to assess the structural and functional defects associated with a large set of 117 distinct BRCA1 missense variants within the essential BRCT domain of the BRCA1 protein that have been documented in individuals with a family history of breast or ovarian cancer. In the first method, we used limited proteolysis to assess the protein folding stability of each of the mutants compared with the wild-type. In the second method, we used a phosphopeptide pull-down assay to assess the ability of each of the variants to specifically interact with a peptide containing a pSer-X-X-Phe motif, a known functional target of the BRCA1 BRCT domain. Finally, we used transcriptional assays to assess the ability of each BRCT variant to act as a transcriptional activation domain in human cells. Through a correlation of the assay results with available family history and clinical data, we define limits to predict the disease risk associated with each variant. Forty-two of the variants show little effect on function and are likely to represent variants with little or no clinical significance; 50 display a clear functional effect and are likely to represent pathogenic variants; and the remaining 25 variants display intermediate activities. The excellent agreement between the structure/function effects of these mutations and available clinical data supports the notion that functional and structure information can be useful in the development of models to assess cancer risk.
Skeletal muscle dysfunction in survivors of pneumonia disproportionately affects older individuals in whom it causes substantial morbidity. We found that skeletal muscle recovery was impaired in old ...compared with young mice after influenza A virus‐induced pneumonia. In young mice, recovery of muscle loss was associated with expansion of tissue‐resident skeletal muscle macrophages and downregulation of MHC II expression, followed by a proliferation of muscle satellite cells. These findings were absent in old mice and in mice deficient in Cx3cr1. Transcriptomic profiling of tissue‐resident skeletal muscle macrophages from old compared with young mice showed downregulation of pathways associated with phagocytosis and proteostasis, and persistent upregulation of inflammatory pathways. Consistently, skeletal muscle macrophages from old mice failed to downregulate MHCII expression during recovery from influenza A virus‐induced pneumonia and showed impaired phagocytic function in vitro. Like old animals, mice deficient in the phagocytic receptor Mertk showed no macrophage expansion, MHCII downregulation, or satellite cell proliferation and failed to recover skeletal muscle function after influenza A pneumonia. Our data suggest that a loss of phagocytic function in a CX3CR1+ tissue‐resident skeletal muscle macrophage population in old mice precludes satellite cell proliferation and recovery of skeletal muscle function after influenza A pneumonia.
A schematic illustrating how impaired phagocytic function in CX3CR1+ tissue‐resident skeletal muscle macrophages prevents muscle recovery after influenza A virus‐induced pneumonia in old mice
Ctp1 (also known as CtIP or Sae2) collaborates with Mre11-Rad50-Nbs1 to initiate repair of DNA double-strand breaks (DSBs), but its functions remain enigmatic. We report that tetrameric ...Schizosaccharomyces pombe Ctp1 contains multivalent DNA-binding and DNA-bridging activities. Through structural and biophysical analyses of the Ctp1 tetramer, we define the salient features of Ctp1 architecture: an N-terminal interlocking tetrameric helical dimer-of-dimers (THDD) domain and a central intrinsically disordered region (IDR) linked to C-terminal 'RHR' DNA-interaction motifs. The THDD, IDR and RHR are required for Ctp1 DNA-bridging activity in vitro, and both the THDD and RHR are required for efficient DSB repair in S. pombe. Our results establish non-nucleolytic roles of Ctp1 in binding and coordination of DSB-repair intermediates and suggest that ablation of human CtIP DNA binding by truncating mutations underlie the CtIP-linked Seckel and Jawad syndromes.
Genomic knowledge is being translated into clinical care. To fully realize the value, it is critical to place credible information in the hands of clinicians in time to support clinical decision ...making. The electronic health record is an essential component of clinician workflow. Utilizing the electronic health record to present information to support the use of genomic medicine in clinical care to improve outcomes represents a tremendous opportunity. However, there are numerous barriers that prevent the effective use of the electronic health record for this purpose. The electronic health record working groups of the Electronic Medical Records and Genomics (eMERGE) Network and the Clinical Genome Resource (ClinGen) project, along with other groups, have been defining these barriers, to allow the development of solutions that can be tested using implementation pilots. In this paper, we present "lessons learned" from these efforts to inform future efforts leading to the development of effective and sustainable solutions that will support the realization of genomic medicine.
The safety and efficacy of Onyx for the embolization of central nervous system (CNS) arteriovenous (AV) lesions have been widely reported in adults. However, data describing the use of this agent in ...children are limited. This study presents our experience with Onyx in the treatment of CNS AV lesions in pediatric patients.
We retrospectively analyzed clinical and imaging records of 15 pediatric patients who underwent 36 transarterial embolizations by using Onyx for CNS AV lesions, from March 2007 through April 2009 at our institution. Underlying pathologies included brain AV malformations (AVMs) (n = 7), vein of Galen malformations (n = 4), dural AV fistulas (n = 2), and spinal AVMs (n = 2). For 7 procedures in very high-flow lesions, detachable coils were deployed before Onyx embolization, whereas in 29 procedures, Onyx was the sole embolic agent. The efficacy of embolization was judged by the residuum of AV shunting within the target region.
Embolization was complete in 2 patients, nearly complete in 9 patients, and partial (and ongoing) in 4 patients. Following staged embolization, 7 patients underwent surgical resection without significant blood loss and with good functional outcome in all cases. Clinically silent non-target embolization was encountered in 2 of 36 procedures. After 3 of the 36 embolizations, patients developed transient neurologic symptoms, all of which resolved to baseline within 24 hours. There were no non-neurologic adverse events. There was no imaging evidence of infarct or hemorrhage.
Onyx embolization of pediatric CNS AV lesions can be an efficacious treatment technique, with extremely low associated morbidity.
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