20 research outputs found
An accessible proteogenomics informatics resource for cancer researchers
Proteogenomics has emerged as a valuable approach in cancer research, which integrates genomic and transcriptomic data with mass spectrometry–based proteomics data to directly identify expressed, variant protein sequences that may have functional roles in cancer. This approach is computationally intensive, requiring integration of disparate software tools into sophisticated workflows, challenging its adoption by nonexpert, bench scientists. To address this need, we have developed an extensible, Galaxy-based resource aimed at providing more researchers access to, and training in, proteogenomic informatics. Our resource brings together software from several leading research groups to address two foundational aspects of proteogenomics: (i) generation of customized, annotated protein sequence databases from RNA-Seq data; and (ii) accurate matching of tandem mass spectrometry data to putative variants, followed by filtering to confirm their novelty. Directions for accessing software tools and workflows, along with instructional documentation, can be found at z.umn.edu/canresgithub.publishedVersio
Induction of 8,5′-Cyclo-2′-deoxyadenosine and 8,5′-Cyclo-2′-deoxyguanosine in Isolated DNA by Fenton-Type Reagents
Exposure of aqueous solutions of
DNA to X- or γ-rays, which
induces the hydroxyl radical as one of the major reactive oxygen species
(ROS), can result in the generation of a battery of single-nucleobase
and bulky DNA lesions. These include the (5′<i>R</i>) and (5′<i>S</i>) diastereomers of 8,5′-cyclo-2′-deoxyadenosine
(cdA) and 8,5′-cyclo-2′-deoxyguanosine (cdG), which
were also found to be present at appreciable levels in DNA isolated
from mammalian cells and tissues. However, it remains unexplored how
efficiently the cdA and cdG can be induced by Fenton-type reagents.
By employing HPLC coupled with tandem mass spectrometry (LC-MS/MS/MS)
with the use of the isotope-dilution technique, here we demonstrated
that treatment of calf thymus DNA with Cu(II) or Fe(II), together
with H<sub>2</sub>O<sub>2</sub> and ascorbate, could lead to dose-responsive
formation of both the (5′<i>R</i>) and (5′<i>S</i>) diastereomers of cdA and cdG, though the yields of cdG
were 2–4 orders of magnitude lower than that of 8-oxo-7,8-dihydro-2′-deoxyguanosine.
This result suggests that the Fenton reaction may constitute an important
endogenous source for the formation of the cdA and cdG. Additionally,
the (5′<i>R</i>) diastereomers of cdA and cdG were
induced at markedly higher levels than the (5′<i>S</i>) counterparts. This latter finding, in conjunction with the previous
observations of similar or greater levels of the (5′<i>S</i>) than (5′<i>R</i>) diastereomers of the
two lesions in mammalian tissues, furnishes an additional line of
evidence to support the more efficient repair of the (5′<i>R</i>) diastereomers of the purine cyclonucleosides in mammalian
cells
Developing A Baseline Metabolomic Signature Associated with COVID-19 Severity: Insights from Prospective Trials Encompassing 13 U.S. Centers
Metabolic disease is a significant risk factor for severe COVID-19 infection, but the contributing pathways are not yet fully elucidated. Using data from two randomized controlled trials across 13 U.S. academic centers, our goal was to characterize metabolic features that predict severe COVID-19 and define a novel baseline metabolomic signature. Individuals (n = 133) were dichotomized as having mild or moderate/severe COVID-19 disease based on the WHO ordinal scale. Blood samples were analyzed using the Biocrates platform, providing 630 targeted metabolites for analysis. Resampling techniques and machine learning models were used to determine metabolomic features associated with severe disease. Ingenuity Pathway Analysis (IPA) was used for functional enrichment analysis. To aid in clinical decision making, we created baseline metabolomics signatures of low-correlated molecules. Multivariable logistic regression models were fit to associate these signatures with severe disease on training data. A three-metabolite signature, lysophosphatidylcholine a C17:0, dihydroceramide (d18:0/24:1), and triacylglyceride (20:4_36:4), resulted in the best discrimination performance with an average test AUROC of 0.978 and F1 score of 0.942. Pathways related to amino acids were significantly enriched from the IPA analyses, and the mitogen-activated protein kinase kinase 5 (MAP2K5) was differentially activated between groups. In conclusion, metabolites related to lipid metabolism efficiently discriminated between mild vs. moderate/severe disease. SDMA and GABA demonstrated the potential to discriminate between these two groups as well. The mitogen-activated protein kinase kinase 5 (MAP2K5) regulator is differentially activated between groups, suggesting further investigation as a potential therapeutic pathway
Unique-region phosphorylation targets LynA for rapid degradation, tuning its expression and signaling in myeloid cells
The activity of Src-family kinases (SFKs), which phosphorylate immunoreceptor tyrosine-based activation motifs (ITAMs), is a critical factor regulating myeloid-cell activation. We reported previously that the SFK LynA is uniquely susceptible to rapid ubiquitin-mediated degradation in macrophages, functioning as a rheostat regulating signaling (Freedman et al., 2015). We now report the mechanism by which LynA is preferentially targeted for degradation and how cell specificity is built into the LynA rheostat. Using genetic, biochemical, and quantitative phosphopeptide analyses, we found that the E3 ubiquitin ligase c-Cbl preferentially targets LynA via a phosphorylated tyrosine (Y32) in its unique region. This distinct mode of c-Cbl recognition depresses steady-state expression of LynA in macrophages derived from mice. Mast cells, however, express little c-Cbl and have correspondingly high LynA. Upon activation, mast-cell LynA is not rapidly degraded, and SFK-mediated signaling is amplified relative to macrophages. Cell-specific c-Cbl expression thus builds cell specificity into the LynA checkpoint
An accessible proteogenomics informatics resource for cancer researchers
Proteogenomics has emerged as a valuable approach in cancer research, which integrates genomic and transcriptomic data with mass spectrometry-based proteomics data to directly identify expressed, variant protein sequences that may have functional roles in cancer. This approach is computationally intensive, requiring integration of disparate software tools into sophisticated workflows, challenging its adoption by nonexpert, bench scientists. To address this need, we have developed an extensible, Galaxy-based resource aimed at providing more researchers access to, and training in, proteogenomic informatics. Our resource brings together software from several leading research groups to address two foundational aspects of proteogenomics: (i) generation of customized, annotated protein sequence databases from RNA-Seq data; and (ii) accurate matching of tandem mass spectrometry data to putative variants, followed by filtering to confirm their novelty. Directions for accessing software tools and workflows, along with instructional documentation, can be found at z.umn.edu/canresgithub. (C) 2017 AACR
An accessible proteogenomics informatics resource for cancer researchers
Proteogenomics has emerged as a valuable approach in cancer research, which integrates genomic and transcriptomic data with mass spectrometry–based proteomics data to directly identify expressed, variant protein sequences that may have functional roles in cancer. This approach is computationally intensive, requiring integration of disparate software tools into sophisticated workflows, challenging its adoption by nonexpert, bench scientists. To address this need, we have developed an extensible, Galaxy-based resource aimed at providing more researchers access to, and training in, proteogenomic informatics. Our resource brings together software from several leading research groups to address two foundational aspects of proteogenomics: (i) generation of customized, annotated protein sequence databases from RNA-Seq data; and (ii) accurate matching of tandem mass spectrometry data to putative variants, followed by filtering to confirm their novelty. Directions for accessing software tools and workflows, along with instructional documentation, can be found at z.umn.edu/canresgithub
Oxidized mitochondrial DNA activates the NLRP3 inflammasome during apoptosis
We report that in the presence of signal 1 (NF-κB), the NLRP3 inflammasome was activated by mitochondrial apoptotic signaling that licensed production of interleukin-1β (IL-1β). NLRP3 secondary signal activators such as ATP induced mitochondrial dysfunction and apoptosis, resulting in release of oxidized mitochondrial DNA (mtDNA) into the cytosol, where it bound to and activated the NLRP3 inflammasome. The antiapoptotic protein Bcl-2 inversely regulated mitochondrial dysfunction and NLRP3 inflammasome activation. Mitochondrial DNA directly induced NLRP3 inflammasome activation, because macrophages lacking mtDNA had severely attenuated IL-1β production, yet still underwent apoptosis. Both binding of oxidized mtDNA to the NLRP3 inflammasome and IL-1β secretion could be competitively inhibited by the oxidized nucleoside 8-OH-dG. Thus, our data reveal that oxidized mtDNA released during programmed cell death causes activation of the NLRP3 inflammasome. These results provide a missing link between apoptosis and inflammasome activation, via binding of cytosolic oxidized mtDNA to the NLRP3 inflammasome
Tet-Mediated Formation of 5‑Hydroxymethylcytosine in RNA
Oxidation of 5-methylcytosine
in DNA by ten-eleven translocation
(Tet) family of enzymes has been demonstrated to play a significant
role in epigenetic regulation in mammals. We found that Tet enzymes
also possess the activity of catalyzing the formation of 5-hydroxymethylcytidine
(5-hmrC) in RNA <i>in vitro</i>. In addition, the catalytic
domains of all three Tet enzymes as well as full-length Tet3 could
induce the formation of 5-hmrC in human cells. Moreover, 5-hmrC was
present at appreciable levels (∼1 per 5000 5-methylcytidine)
in RNA of mammalian cells and tissues. Our results suggest the involvement
of this oxidation in RNA biology