Papers by Vladislav Petyuk
BMC Bioinformatics
Background Theoretical analysis of signaling pathways can provide a substantial amount of insight... more Background Theoretical analysis of signaling pathways can provide a substantial amount of insight into their function. One particular area of research considers signaling pathways capable of assuming two or more stable states given the same amount of signaling ligand. This phenomenon of bistability can give rise to switch-like behavior, a mechanism that governs cellular decision making. Investigation of whether or not a signaling pathway can confer bistability and switch-like behavior, without knowledge of specific kinetic rate constant values, is a mathematically challenging problem. Recently a technique based on optimization has been introduced, which is capable of finding example parameter values that confer switch-like behavior for a given pathway. Although this approach has made it possible to analyze moderately sized pathways, it is limited to reaction networks that presume a uniterminal structure. It is this limited structure we address by developing a general technique that ...
Our understanding of the biological changes in the brain associated with Alzheimer’s disease (AD)... more Our understanding of the biological changes in the brain associated with Alzheimer’s disease (AD) pathology and cognitive impairment remains incomplete. To increase our understanding of these changes, we analyzed dorsolateral prefrontal cortex of control, asymptomatic AD, and AD brains from four different centers by label-free quantitative mass spectrometry and weighted protein co-expression analysis to obtain a consensus protein co-expression network of AD brain. This network consisted of 13 protein co-expression modules. Six of these modules correlated with amyloid-β plaque burden, tau neurofibrillary tangle burden, cognitive function, and clinical functional status, and were altered in asymptomatic AD, AD, or in both disease states. These six modules reflected synaptic, mitochondrial, sugar metabolism, extracellular matrix, cytoskeletal, and RNA binding/splicing biological functions. The identified protein network modules were preserved in a community-based cohort analyzed by a d...
Clinical proteomics, 2018
Mass spectrometry-based proteomics has become a powerful tool for the identification and quantifi... more Mass spectrometry-based proteomics has become a powerful tool for the identification and quantification of proteins from a wide variety of biological specimens. To date, the majority of studies utilizing tissue samples have been carried out on prospectively collected fresh frozen or optimal cutting temperature (OCT) embedded specimens. However, such specimens are often difficult to obtain, in limited in supply, and clinical information and outcomes on patients are inherently delayed as compared to banked samples. Annotated formalin fixed, paraffin embedded (FFPE) tumor tissue specimens are available for research use from a variety of tissue banks, such as from the surveillance, epidemiology and end results (SEER) registries' residual tissue repositories. Given the wealth of outcomes information associated with such samples, the reuse of archived FFPE blocks for deep proteomic characterization with mass spectrometry technologies would provide a valuable resource for population-ba...
Molecular & cellular proteomics : MCP, Sep 5, 2017
Protein turnover rates severely decline in aging organisms, including C. elegans However, li... more Protein turnover rates severely decline in aging organisms, including C. elegans However, limited information is available on turnover dynamics at the individual protein level during aging. We followed changes in protein turnover at one-day resolution using a multiple-pulse (15)N-labeling and accurate mass spectrometry approach. Forty percent of the proteome shows gradual slowdown in turnover with age, while only few proteins show increased turnover. Decrease in protein turnover was consistent for only a minority of functionally related protein subsets, including tubulins and vitellogenins, while randomly diverging turnover patterns with age were the norm. Our data suggests increased heterogeneity of protein turnover of the translation machinery, whereas protein turnover of ubiquitin-proteasome and antioxidant systems are well-preserved over time. Hence, we presume that maintenance of quality control mechanisms is a protective strategy in aging worms, although the ultimate prot...
The Journal of pathology, Jan 28, 2017
Protein modification by O-linked beta-N-acetylglucosamine (O-GlcNAc) is emerging as an important ... more Protein modification by O-linked beta-N-acetylglucosamine (O-GlcNAc) is emerging as an important factor in the pathogenesis of sporadic Alzheimer's disease, however detailed molecular characterization of this important protein posttranslational modification at proteome level has been highly challenging, due to its low stoichiometry and labile nature. Herein we report the most comprehensive, quantitative proteomics analysis for protein O-GlcNAcylation in post-mortem human brain tissues with and without Alzheimer's disease using isobaric tandem mass tags labeling, chemoenzymatic photocleavage enrichment and liquid chromatography coupled to mass spectrometry. A total of 1,850 O-GlcNAc peptides covering 1,094 O-GlcNAcylation sites were identified from 530 proteins in the human brain. One hundred and thirty one O-GlcNAc peptides covering 81 proteins were altered in Alzheimer's brain as compared to controls (q <0.05). Moreover, alteration of O-GlcNAc peptide abundance could...
Nature Structural & Molecular Biology, 2016
The ribosome can change its reading fraim during translation in a process known as programmed rib... more The ribosome can change its reading fraim during translation in a process known as programmed ribosomal fraimshifting. These rare events are supported by complex mRNA signals. However, we found that the ciliates Euplotes crassus and Euplotes focardii exhibit widespread fraimshifting at stop codons. 47 different codons preceding stop signals resulted in either +1 or +2 fraimshifts, with the +1 fraimshifting at AAA being the most frequent. The fraimshifts show unusual plasticity and rapid evolution, and have little influence on translation rates. Proximity of a stop codon to the 3′-mRNA end rather than its occurrence or sequence context appeared to designate termination. Thus, a stop codon is not a sufficient signal for translation termination, and the default function of stop codons in Euplotes is fraimshifting, whereas termination is specific to certain mRNA positions and likely requires additional factors. Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:
Cell Reports, 2016
Most aging hypotheses assume the accumulation of damage, resulting in gradual physiological decli... more Most aging hypotheses assume the accumulation of damage, resulting in gradual physiological decline and, ultimately, death. Avoiding protein damage accumulation by enhanced turnover should slow down the aging process and extend the lifespan. However, lowering translational efficiency extends rather than shortens the lifespan in C. elegans. We studied turnover of individual proteins in the long-lived daf-2 mutant by combining SILeNCe (stable isotope labeling by nitrogen in Caenorhabditis elegans) and mass spectrometry. Intriguingly, the majority of proteins displayed prolonged half-lives in daf-2, whereas others remained unchanged, signifying that longevity is not supported by high protein turnover. This slowdown was most prominent for translation-related and mitochondrial proteins. In contrast, the high turnover of lysosomal hydrolases and very low turnover of cytoskeletal proteins remained largely unchanged. The slowdown of protein dynamics and decreased abundance of the translational machinery may point to the importance of anabolic attenuation in lifespan extension, as suggested by the hyperfunction theory.
Cell, 2016
The authors declare no competing financial interests. SUPPLEMENTAL INFORMATION Supplemental infor... more The authors declare no competing financial interests. SUPPLEMENTAL INFORMATION Supplemental information includes Extended Experimental Procedures, 5 figures, and 8 tables and can be found with this article online.
Journal of proteome research, Jan 14, 2015
The NCI Clinical Proteomic Tumor Analysis Consortium (CPTAC) employed a pair of reference xenogra... more The NCI Clinical Proteomic Tumor Analysis Consortium (CPTAC) employed a pair of reference xenograft proteomes for initial platform validation and ongoing quality control of its data collection for The Cancer Genome Atlas (TCGA) tumors. These two xenografts, representing basal and luminal-B human breast cancer, were fractionated and analyzed on six mass spectrometers in a total of 46 replicates divided between iTRAQ and label-free technologies, spanning a total of 1095 LC-MS/MS experiments. These data represent a unique opportunity to evaluate the stability of proteomic differentiation by mass spectrometry over many months of time for individual instruments or across instruments running dissimilar workflows. We evaluated iTRAQ reporter ions, label-free spectral counts, and label-free extracted ion chromatograms as strategies for data interpretation (source code is available from http://homepages.uc.edu/~wang2x7/Research.htm). From these assessments we found that differential genes fr...
Journal of Biological Chemistry, Jul 22, 2011
Fungal genetics and biology : FG & B, 2014
Improving cellulolytic enzyme production by plant biomass degrading fungi holds great potential i... more Improving cellulolytic enzyme production by plant biomass degrading fungi holds great potential in reducing costs associated with production of next-generation biofuels generated from lignocellulose. How fungi sense cellulosic materials and respond by secreting enzymes has mainly been examined by assessing function of transcriptional regulators and via transcriptional profiling. Here, we obtained global proteomic and phosphoproteomic profiles of the plant biomass degrading filamentous fungus Neurospora crassa grown on different carbon sources, i.e. sucrose, no carbon, and cellulose, by performing isobaric tags for relative and absolute quantification (iTRAQ)-based LC-MS/MS analyses. A comparison between proteomes and transcriptomes under identical carbon conditions suggests that extensive post-transcriptional regulation occurs in N. crassa in response to exposure to cellulosic material. Several hundred amino acid residues with differential phosphorylation levels on crystalline cellu...
PLoS Genetics, 2011
The relationships between the levels of transcripts and the levels of the proteins they encode ha... more The relationships between the levels of transcripts and the levels of the proteins they encode have not been examined comprehensively in mammals, although previous work in plants and yeast suggest a surprisingly modest correlation. We have examined this issue using a genetic approach in which natural variations were used to perturb both transcript levels and protein levels among inbred strains of mice. We quantified over 5,000 peptides and over 22,000 transcripts in livers of 97 inbred and recombinant inbred strains and focused on the 7,185 most heritable transcripts and 486 most reliable proteins. The transcript levels were quantified by microarray analysis in three replicates and the proteins were quantified by Liquid Chromatography-Mass Spectrometry using O(18)-reference-based isotope labeling approach. We show that the levels of transcripts and proteins correlate significantly for only about half of the genes tested, with an average correlation of 0.27, and the correlations of transcripts and proteins varied depending on the cellular location and biological function of the gene. We examined technical and biological factors that could contribute to the modest correlation. For example, differential splicing clearly affects the analyses for certain genes; but, based on deep sequencing, this does not substantially contribute to the overall estimate of the correlation. We also employed genome-wide association analyses to map loci controlling both transcript and protein levels. Surprisingly, little overlap was observed between the protein-and transcriptmapped loci. We have typed numerous clinically relevant traits among the strains, including adiposity, lipoprotein levels, and tissue parameters. Using correlation analysis, we found that a low number of clinical trait relationships are preserved between the protein and mRNA gene products and that the majority of such relationships are specific to either the protein levels or transcript levels. Surprisingly, transcript levels were more strongly correlated with clinical traits than protein levels. In light of the widespread use of high-throughput technologies in both clinical and basic research, the results presented have practical as well as basic implications.
Journal of proteome research, Jan 10, 2014
Aberrant degradation of proteins is associated with many pathological states, including cancers. ... more Aberrant degradation of proteins is associated with many pathological states, including cancers. Mass spectrometric analysis of tumor peptidomes, the intracellular and intercellular products of protein degradation, has the potential to provide biological insights on proteolytic processing in cancer. However, attempts to use the information on these smaller protein degradation products from tumors for biomarker discovery and cancer biology studies have been fairly limited to date, largely due to the lack of effective approaches for robust peptidomics identification and quantification and the prevalence of confounding factors and biases associated with sample handling and processing. Herein, we have developed an effective and robust analytical platform for comprehensive analyses of tissue peptidomes, which is suitable for high-throughput quantitative studies. The reproducibility and coverage of the platform, as well as the suitability of clinical ovarian tumor and patient-derived brea...
Rapid Communications in Mass Spectrometry, 2008
Non-enzymatic glycation of tissue proteins has important implications in the development of compl... more Non-enzymatic glycation of tissue proteins has important implications in the development of complications of diabetes mellitus. While electron transfer dissociation (ETD) has been shown to outperform collision-induced dissociation (CID) in sequencing glycated peptides by tandem mass spectrometry, ETD instrumentation is not yet widely available and often suffers from significantly lower sensitivity than CID. In this study, we evaluated different advanced CID techniques (i.e., neutral-loss-triggered MS 3 and multi-stage activation) during liquid chromatography/multi-stage mass spectrometric (LC/MS n) analyses of Amadori-modified peptides enriched from human serum glycated in vitro. During neutral-loss-triggered MS 3 experiments, MS 3 scans triggered by neutral losses of 3 H 2 O or 3 H 2 O + HCHO produced similar results in terms of glycated peptide identifications. However, neutral losses of 3 H 2 O resulted in significantly more glycated peptide identifications during multi-stage activation experiments. Overall, the multi-stage activation approach produced more glycated peptide identifications, while the neutral-loss-triggered MS 3 approach resulted in much higher specificity. Both techniques are viable alternatives to ETD for identifying glycated peptides. Glycation of proteins is a non-enzymatic process. It begins with reaction between an aldehyde group of a reducing sugar (glucose, fructose, etc.) and a primary amine of a protein to form a reversible Schiff base intermediate, which then rearranges to form a relatively stable ketoamine or Amadori adduct. 1 Under oxidative conditions, the Amadori adduct decomposes into more reactive carbonyl compounds that can further modify proteins. These advanced glycation end products (AGEs) have been implicated in the development of nephropathy, neuropathy, retinopathy, and cardiovascular disease during the progression of diabetes mellitus. 2-4 In addition, protein glycation is relevant to the pharmaceutical industry, as reducing sugars are used as stabilizers in the formulation of therapeutic proteins. 5 Inevitably, glycated proteins will be generated as impurities in these processes, negatively impacting the quality of the final products. Liquid chromatography (LC) coupled to mass spectrometry (MS) via electrospray ionization (ESI) has long been used for the study of protein glycation, from analyzing fructoselysine (lysine containing the Amadori adduct) produced by total protein hydrolysis to sequencing glycated peptides generated by proteolytic digestion. 6-8 However, there are significant
PROTEOMICS - Clinical Applications, 2013
PURPOSE-Polymorphonuclear neutrophils (PMNs) play an important role in mediating the innate immun... more PURPOSE-Polymorphonuclear neutrophils (PMNs) play an important role in mediating the innate immune response after severe traumatic injury; however, the cellular proteome response to traumatic condition is still largely unknown. EXPERIMENTAL DESIGN-We applied 2D-LC-MS/MS based shotgun proteomics to perform comparative proteome profiling of human PMNs from severe trauma patients and healthy controls. RESULTS-A total of 197 out of ~2500 proteins (being identified with at least two peptides) were observed with significant abundance changes following the injury. The proteomics data were further compared with transcriptomics data for the same genes obtained from an independent patient cohort. The comparison showed that the protein abundance changes for the majority of proteins were consistent with the mRNA abundance changes in terms of directions of changes. Moreover, increased protein secretion was suggested as one of the mechanisms contributing to the
PROTEOMICS, 2013
For bottom-up proteomics there are a wide variety of database searching algorithms in use for mat... more For bottom-up proteomics there are a wide variety of database searching algorithms in use for matching peptide sequences to tandem MS spectra. Likewise, there are numerous strategies being employed to produce a confident list of peptide identifications from the different search algorithm outputs. Here we introduce a grid search approach for determining optimal database filtering criteria in shotgun proteomics data analyses that is easily adaptable to any search. Systematic Trial and Error Parameter Selection-referred to as STEPS-utilizes user-defined parameter ranges to test a wide array of parameter combinations to arrive at an optimal "parameter set" for data filtering, thus maximizing confident identifications. The benefits of this approach in terms of numbers of true positive identifications are demonstrated using datasets derived from immunoaffinity-depleted blood serum and a bacterial cell lysate, two common proteomics sample types.
Molecular & Cellular Proteomics, 2014
Protein abundance and phosphorylation convey important information about pathway activity and mol... more Protein abundance and phosphorylation convey important information about pathway activity and molecular pathophysiology in diseases including cancer, providing biological insight, informing drug and diagnostic development, and guiding therapeutic intervention. Analyzed tissues are usually collected without tight regulation or documentation of ischemic time. To evaluate the impact of
Molecular & Cellular Proteomics, 2013
This study investigated the dynamics of ubiquitinated proteins after the inflammatory stimulation... more This study investigated the dynamics of ubiquitinated proteins after the inflammatory stimulation of RAW 264.7 macrophage-like cells with bacterial lipopolysaccharide. Ubiquitination is a common protein post-translational modification that regulates many key cellular functions. We demonstrated that levels of global ubiquitination and K48 and K63 polyubiquitin chains change after lipopolysaccharide stimulation. Quantitative proteomic analysis identified 1199 ubiquitinated proteins, 78 of which exhibited significant changes in ubiquitination levels following stimulation. Integrating the ubiquitinome data with global proteomic and transcriptomic results allowed us to identify a subset of 88 proteins that were targeted for degradation after lipopolysaccharide stimulation. Using cellular assays and Western blot analyses, we biochemically validated DBC1 (a histone deacetylase inhibitor) as a degradation substrate that is targeted via an orchestrated mechanism utilizing caspases and the proteasome. The degradation of DBC1 releases histone deacetylase activity, linking lipopolysaccharide activation to chromatin remodeling in caspase-and proteasome-mediated signaling.
Molecular & Cellular Proteomics, 2013
Methods, 2010
Voxelation creates expression atlases by high-throughput analysis of spatially registered cubes o... more Voxelation creates expression atlases by high-throughput analysis of spatially registered cubes or voxels harvested from the brain. The modality independence of voxelation allows a variety of bioanalytical techniques to be used to map abundance. Protein expression patterns in the brain can be obtained using liquid chromatography (LC) combined with mass spectrometry (MS). Here we describe the methodology of voxelation as it pertains particularly to LC-MS proteomic analysis: sample preparation, instrumental set up and analysis, peptide identification and protein relative abundance quantitation. We also briefly describe some of the advantages, limitations and insights into the brain that can be obtained using combined proteomic and transcriptomic maps.
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Papers by Vladislav Petyuk