The large number of genomes that will be sequenced will need to be annotated with genes and other functional features. Aligning gene sequences from a related species to the target genome is an ...economical and highly reliable method to identify genes; unfortunately, existing tools have been lacking in sensitivity and speed. A program we reported, sim4cc, was shown to be highly accurate but is limited to comparing one cDNA with one genomic sequence. We present here an optimization of the tool, implemented in the packages sim4db and leaff. The new tool performs batch alignments of cDNA and genomic sequences in a fraction of the time required by its predecessor, and thus is very well suited for genome-wide analyses.
Availability:
Sim4db and leaff are written in C, C++ and Perl for Linux and other Unix platforms. Source code is distributed free of charge from http://sourceforge.net/projects/kmer/.
Contact:
florea@umiacs.umd.edu
Supplementary information:
Supplementary data are available at Bioinformatics Online.
Alternative splicing is widely recognized for its roles in regulating genes and creating gene diversity. However, despite many efforts, the repertoire of gene splicing variation is still incompletely ...characterized, even in humans. Here we describe a new computational system, ASprofile, and its application to RNA-seq data from Illumina's Human Body Map project (>2.5 billion reads). Using the system, we identified putative alternative splicing events in 16 different human tissues, which provide a dynamic picture of splicing variation across the tissues. We detected 26,989 potential exon skipping events representing differences in splicing patterns among the tissues. A large proportion of the events (>60%) were novel, involving new exons (~3000), new introns (~16000), or both. When tracing these events across the sixteen tissues, only a small number (4-7%) appeared to be differentially expressed ('switched') between two tissues, while 30-45% showed little variation, and the remaining 50-65% were not present in one or both tissues compared. Novel exon skipping events appeared to be slightly less variable than known events, but were more tissue-specific. Our study represents the first effort to build a comprehensive catalog of alternative splicing in normal human tissues from RNA-seq data, while providing insights into the role of alternative splicing in shaping tissue transcriptome differences. The catalog of events and the ASprofile software are freely available from the Zenodo repository ( http://zenodo.org/record/7068; doi: 10.5281/zenodo.7068) and from our web site http://ccb.jhu.edu/software/ASprofile.
BACKGROUNDGermline mutations in telomerase and other telomere maintenance genes manifest in the premature aging short telomere syndromes. Myelodysplastic syndromes and acute myeloid leukemia ...(MDS/AML) account for 75% of associated malignancies, but how these cancers overcome the inherited telomere defect is unknown.METHODSWe used ultra-deep targeted sequencing to detect somatic reversion mutations in 17 candidate telomere lengthening genes among controls and patients with short telomere syndromes with and without MDS/AML, and we tested the functional significance of these mutations.RESULTSWhile no controls carried somatic mutations in telomere maintenance genes, 29% (16 of 56) of adults with germline telomere maintenance defects carried at least 1 (P < 0.001), and 13% (7 of 56) had 2 or more. In addition to TERT promoter mutations, which were present in 19%, another 13% of patients carried a mutation in POT1 or TERF2IP. POT1 mutations impaired telomere binding in vitro and some mutations were identical to ones seen in familial melanoma associated with longer telomere length. Exclusively in patients with germline defects in telomerase RNA (TR), we identified somatic mutations in nuclear RNA exosome genes RBM7, SKIV2L2, and DIS3, where loss-of-function upregulates mature TR levels. Somatic reversion events in 6 telomere-related genes were more prevalent in patients who were MDS/AML-free (P = 0.02, RR 4.4, 95% CI 1.2-16.7), and no patient with MDS/AML had more than 1 reversion mutation.CONCLUSIONOur data indicate that diverse adaptive somatic mutations arise in the short telomere syndromes. Their presence may alleviate the telomere crisis that promotes transformation to MDS/AML.FUNDINGThis work was supported by the NIH, the Commonwealth Foundation, the S&R Foundation Kuno Award, the Williams Foundation, the Vera and Joseph Dresner Foundation, the MacMillan Pathway to Independence Award, the American Society of Hematology Scholar Award, the Johns Hopkins Research Program for Medical Students, and the Turock Scholars Fund.
Heart failure is a leading cause of death worldwide and is associated with the rising prevalence of obesity, hypertension, and diabetes.
-GlcNAcylation (the attachment of
-linked ...β-N-acetylglucosamine
-GlcNAc moieties to cytoplasmic, nuclear, and mitochondrial proteins) is a posttranslational modification of intracellular proteins and serves as a metabolic rheostat for cellular stress. Total levels of
-GlcNAcylation are determined by nutrient and metabolic flux, in addition to the net activity of 2 enzymes:
-GlcNAc transferase (OGT) and
-GlcNAcase (OGA). Failing myocardium is marked by increased
-GlcNAcylation, but whether excessive
-GlcNAcylation contributes to cardiomyopathy and heart failure is unknown.
We developed 2 new transgenic mouse models with myocardial overexpression of OGT and OGA to control
-GlcNAcylation independent of pathologic stress.
We found that OGT transgenic hearts showed increased
-GlcNAcylation and developed severe dilated cardiomyopathy, ventricular arrhythmias, and premature death. In contrast, OGA transgenic hearts had lower
-GlcNAcylation but identical cardiac function to wild-type littermate controls. OGA transgenic hearts were resistant to pathologic stress induced by pressure overload with attenuated myocardial
-GlcNAcylation levels after stress and decreased pathologic hypertrophy compared with wild-type controls. Interbreeding OGT with OGA transgenic mice rescued cardiomyopathy and premature death, despite persistent elevation of myocardial OGT. Transcriptomic and functional studies revealed disrupted mitochondrial energetics with impairment of complex I activity in hearts from OGT transgenic mice. Complex I activity was rescued by OGA transgenic interbreeding, suggesting an important role for mitochondrial complex I in
-GlcNAc-mediated cardiac pathology.
Our data provide evidence that excessive
-GlcNAcylation causes cardiomyopathy, at least in part, attributable to defective energetics. Enhanced OGA activity is well tolerated and attenuation of
-GlcNAcylation is beneficial against pressure overload-induced pathologic remodeling and heart failure. These findings suggest that attenuation of excessive
-GlcNAcylation may represent a novel therapeutic approach for cardiomyopathy.
Background
Gut microbiota likely impact obesity and metabolic diseases. We evaluated the changes in gut microbiota after surgical versus medical weight loss in adults with diabetes and obesity.
...Methods
We performed 16S rRNA amplicon sequencing to identify the gut microbial composition at baseline and at 10% weight loss in adults with diabetes who were randomized to medical weight loss (MWL,
n
= 4), adjustable gastric banding (AGB,
n
= 4), or Roux-en-Y gastric bypass (RYGB,
n
= 4).
Results
All participants were female, 75% reported black race with mean age of 51 years. At similar weight loss amount and glycemic improvement, the RYGB group had the most number of bacterial species (10 increased, 1 decreased) that significantly changed (
p
< 0.05) in relative abundance. Alpha-diversity at follow-up was significantly lower in AGB group compared to MWL and RYGB (observed species for AGB vs. MWL,
p
= 0.0093; AGB vs. RYGB,
p
= 0.0093). The relative abundance of
Faecalibacterium prausnitzii
increased in 3 participants after RYGB, 1 after AGB, and 1 after MWL.
Conclusions
At similar weight loss and glycemic improvement, the greatest alteration in gut microbiota occurred after RYGB with an increase in the potentially beneficial bacterium,
F. prausnitzii
. Gut microbial diversity tended to decrease after AGB and increase after RYGB and MWL. Future studies are needed to determine the impact and durability of gut microbial changes over time and their role in long-term metabolic improvement after bariatric surgery in adults with type 2 diabetes.
Clinical Trial Registration
NCTDK089557—
ClinicalTrials.gov
Although lung cancer is generally thought to be environmentally provoked, anecdotal familial clustering has been reported, suggesting that there may be genetic susceptibility factors. We ...systematically tested whether germline mutations in eight candidate genes may be risk factors for lung adenocarcinoma.
We studied lung adenocarcinoma cases for which germline sequence data had been generated as part of The Cancer Genome Atlas project but had not been previously analyzed. We selected eight genes, ATM serine/threonine kinase gene (ATM), BRCA2, DNA repair associated gene (BRCA2), checkpoint kinase 2 gene (CHEK2), EGFR, parkin RBR E3 ubiquitin protein ligase gene (PARK2), telomerase reverse transcriptase gene (TERT), tumor protein p53 gene (TP53), and Yes associated protein 1 gene (YAP1), on the basis of prior anecdotal association with lung cancer or genome-wide association studies.
Among 555 lung adenocarcinoma cases, we detected 14 pathogenic mutations in five genes; they occurred at a frequency of 2.5% and represented an OR of 66 (95% confidence interval: 33–125, p < 0.0001 chi-square test). The mutations fell most commonly in ATM (50%), followed by TP53, BRCA2, EGFR, and PARK2. Most (86%) of these variants had been reported in other familial cancer syndromes. Another 12 cases (2%) carried ultrarare variants that were predicted to be deleterious by three protein prediction programs; these most frequently involved ATM and BRCA2.
A subset of patients with lung adenocarcinoma, at least 2.5% to 4.5%, carry germline variants that have been linked to cancer risk in Mendelian syndromes. The genes fall most frequently in DNA repair pathways. Our data indicate that patients with lung adenocarcinoma, similar to other solid tumors, include a subset of patients with inherited susceptibility.
The genome of the domestic cow, Bos taurus, was sequenced using a mixture of hierarchical and whole-genome shotgun sequencing methods.
We have assembled the 35 million sequence reads and applied a ...variety of assembly improvement techniques, creating an assembly of 2.86 billion base pairs that has multiple improvements over previous assemblies: it is more complete, covering more of the genome; thousands of gaps have been closed; many erroneous inversions, deletions, and translocations have been corrected; and thousands of single-nucleotide errors have been corrected. Our evaluation using independent metrics demonstrates that the resulting assembly is substantially more accurate and complete than alternative versions.
By using independent mapping data and conserved synteny between the cow and human genomes, we were able to construct an assembly with excellent large-scale contiguity in which a large majority (approximately 91%) of the genome has been placed onto the 30 B. taurus chromosomes. We constructed a new cow-human synteny map that expands upon previous maps. We also identified for the first time a portion of the B. taurus Y chromosome.
Major histocompatibility complex class I molecules present peptides of 8-10 residues to CD8+ T cells. We used 19 predicted proteomes to determine the influence of CD8+ T cell immune surveillance on ...protein evolution in humans and microbial pathogens by predicting immunopeptidomes, i.e., sets of class I binding peptides present in proteomes. We find that class I peptide binding specificities (i) have had little, if any, influence on the evolution of immunopeptidomes and (ii) do not take advantage of biases in amino acid distribution in proteins other than the concentration of hydrophobic residues in NH2-terminal leader sequences.
Dendritic cells bridge the innate and adaptive immune responses by serving as sensors of infection and as the primary APCs responsible for the initiation of the T cell response against invading ...pathogens. The naive T cell activation requires the following three key signals to be delivered from dendritic cells: engagement of the TCR by peptide Ags bound to MHC molecules (signal 1), engagement of costimulatory molecules on both cell types (signal 2), and expression of polarizing cytokines (signal 3). Initial interactions between Borrelia burgdorferi, the causative agent of Lyme disease, and dendritic cells remain largely unexplored. To address this gap in knowledge, we cultured live B. burgdorferi with monocyte-derived dendritic cells (mo-DCs) from healthy donors to examine the bacterial immunopeptidome associated with HLA-DR. In parallel, we examined changes in the expression of key costimulatory and regulatory molecules as well as profiled the cytokines released by dendritic cells when exposed to live spirochetes. RNA-sequencing studies on B. burgdorferi-pulsed dendritic cells show a unique gene expression signature associated with B. burgdorferi stimulation that differs from stimulation with lipoteichoic acid, a TLR2 agonist. These studies revealed that exposure of mo-DCs to live B. burgdorferi drives the expression of both pro- and anti-inflammatory cytokines as well as immunoregulatory molecules (e.g., PD-L1, IDO1, Tim3). Collectively, these studies indicate that the interaction of live B. burgdorferi with mo-DCs promotes a unique mature DC phenotype that likely impacts the nature of the adaptive T cell response generated in human Lyme disease.
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