The etiology of dental caries is multifactorial, but frequent consumption of free sugars, notably sucrose, appears to be a major factor driving the supragingival microbiota in the direction of dysbiosis. Recent 16S rRNA-based studies indicated that caries-associated communities were less diverse than healthy supragingival plaque but still displayed considerable taxonomic diversity between individuals. Metagenomic studies likewise have found that healthy oral sites from different people were broadly similar with respect to gene function, even though there was an extensive individual variation in their taxonomic profiles. That pattern may also extend to dysbiotic communities. In that case, shifts in community-wide protein relative abundance might provide better biomarkers of dysbiosis that can be achieved through taxonomy alone. In this study, we used a paired oral microcosm biofilm model of dental caries to investigate differences in community composition and protein relative abundance in the presence and absence of sucrose. This approach provided large quantities of protein, which facilitated deep metaproteomic analysis. Community composition was evaluated using 16S rRNA sequencing and metaproteomic approaches. Although taxonomic diversity was reduced by sucrose pulsing, considerable inter-subject variation in community composition remained. By contrast, functional analysis using the SEED ontology found that sucrose induced changes in protein relative abundance patterns for pathways involving glycolysis, lactate production, aciduricity, and ammonia/glutamate metabolism that were conserved across taxonomically diverse dysbiotic oral microcosm biofilm communities. Our findings support the concept of using function-based changes in protein relative abundance as indicators of dysbiosis. Our microcosm model cannot replicate all aspects of the oral environment, but the deep level of metaproteomic analysis it allows makes it suitable for discovering which proteins are most consistently abundant during dysbiosis. It then may be possible to define biomarkers that could be used to detect at-risk tooth surfaces before the development of overt carious lesions.

Background: Ovarian cancer is the most lethal gynecologic malignancy, with the majority of cases diagnosed at an advanced stage when treatments are less successful. Novel serum protein markers are needed to detect ovarian cancer in its earliest stage; when detected early, survival rates are over 90%. The identification of new serum biomarkers is hindered by the presence of a small number of highly abundant proteins that comprise approximately 95% of serum total protein. In this study, we used pooled serum depleted of the most highly abundant proteins to reduce the dynamic range of proteins, and thereby enhance the identification of serum biomarkers using the quantitative proteomic method iTRAQ^®.Results: Medium and low abundance proteins from 6 serum pools of 10 patients each from women with serous ovarian carcinoma, and 6 non-cancer control pools were labeled with isobaric tags using iTRAQ^®to determine the relative abundance of serum proteins identified by MS. A total of 220 unique proteins were identified and fourteen proteins were elevated in ovarian cancer compared to control serum pools, including several novel candidate ovarian cancer biomarkers: extracellular matrix protein-1, leucine-rich alpha-2 glycoprotein-1, lipopolysaccharide binding protein-1, and proteoglycan-4. Western immunoblotting validated the relative increases in serum protein levels for several of the proteins identified.Conclusions: This study provides the first analysis of immunodepleted serum in combination with iTRAQ^®to measure relative protein expression in ovarian cancer patients for the pursuit of serum biomarkers. Several candidate biomarkers were identified which warrant further development.

Background: Two genotypically and microbiologically distinct strains of Mycobacterium avium subsp. paratuberculosis (MAP) exist - S and C MAP strains that primarily infect sheep and cattle, respectively. Concentration of iron in the cultivation medium has been suggested as one contributing factor for the observed microbiologic differences. We recently demonstrated that S strains have defective iron storage systems, leading us to propose that these strains might experience iron toxicity when excess iron is provided in the medium. To test this hypothesis, we carried out transcriptional and proteomic profiling of these MAP strains under iron-replete or -deplete conditions. Results: We first complemented M. smegmatis ΔideR with IdeR of C MAP or that derived from S MAP and compared their transcription profiles using M. smegmatis mc ² 155 microarrays. In the presence of iron, sIdeR repressed expression of bfrA and MAP2073c, a ferritin domain containing protein suggesting that transcriptional control of iron storage may be defective in S strain. We next performed transcriptional and proteomic profiling of the two strain types of MAP under iron-deplete and -replete conditions. Under iron-replete conditions, C strain upregulated iron storage (BfrA), virulence associated (Esx-5 and antigen85 complex), and ribosomal proteins. In striking contrast, S strain downregulated these proteins under iron-replete conditions. iTRAQ (isobaric tag for relative and absolute quantitation) based protein quantitation resulted in the identification of four unannotated proteins. Two of these were upregulated by a C MAP strain in response to iron supplementation. The iron-sparing response to iron limitation was unique to the C strain as evidenced by repression of non-essential iron utilization enzymes (aconitase and succinate dehydrogenase) and upregulation of proteins of essential function (iron transport, [Fe-S] cluster biogenesis and cell division). Conclusions: Taken together, our study revealed that C and S strains of MAP utilize divergent metabolic pathways to accommodate in vitro iron stress. The knowledge of the metabolic pathways these divergent responses play a role in are important to 1) advance our ability to culture the two different strains of MAP efficiently, 2) aid in diagnosis and control of Johne's disease, and 3) advance our understanding of MAP virulence.

Metabolic remodeling plays an important role in the pathophysiology of heart failure (HF). We sought to characterize metabolic remodeling and implicated signaling pathways in two rat models of early systolic dysfunction (MOD), and overt systolic HF (SHF). Tandem mass tag-labeled shotgun proteomics, phospho-(p)-proteomics, and non-targeted metabolomics analyses were performed in left ventricular myocardium tissue from Sham, MOD, and SHF using liquid chromatography–mass spectrometry, n = 3 biological samples per group. Mitochondrial proteins were predominantly down-regulated in MOD (125) and SHF (328) vs. Sham. Of these, 82% (103/125) and 66% (218/328) were involved in metabolism and respiration. Oxidative phosphorylation, mitochondrial fatty acid β-oxidation, Krebs cycle, branched-chain amino acids, and amino acid (glutamine and tryptophan) degradation were highly enriched metabolic pathways that decreased in SHF > MOD. Glycogen and glucose degradation increased predominantly in MOD, whereas glycolysis and pyruvate metabolism decreased predominantly in SHF. PKA signaling at the endoplasmic reticulum–mt interface was attenuated in MOD, whereas overall PKA and AMPK cellular signaling were attenuated in SHF vs. Sham. In conclusion, metabolic remodeling plays an important role in myocardial remodeling. PKA and AMPK signaling crosstalk governs metabolic remodeling in progression to SHF.