Dizeez - main game interface.

<p>Dizeez - main game interface.</p

Concordance between Dizeez-mined associations and Disease and Gene Annotations database.

<p>The ‘concordance ratio’ on the vertical axis is the ratio between the associations supported by DGA and the total number of associations for a given number of votes. ‘7+’ indicates the sum of associations collected with a number of votes between 7 and 11.</p

Quantitative Metaproteomics and Activity-Based Probe Enrichment Reveals Significant Alterations in Protein Expression from a Mouse Model of Inflammatory Bowel Disease

Tandem mass spectrometry based shotgun proteomics of distal gut microbiomes is exceedingly difficult due to the inherent complexity and taxonomic diversity of the samples. We introduce two new methodologies to improve metaproteomic studies of microbiome samples. These methods include the stable isotope labeling in mammals to permit protein quantitation across two mouse cohorts as well as the application of activity-based probes to enrich and analyze both host and microbial proteins with specific functionalities. We used these technologies to study the microbiota from the adoptive T cell transfer mouse model of inflammatory bowel disease (IBD) and compare these samples to an isogenic control, thereby limiting genetic and environmental variables that influence microbiome composition. The data generated highlight quantitative alterations in both host and microbial proteins due to intestinal inflammation and corroborates the observed phylogenetic changes in bacteria that accompany IBD in humans and mouse models. The combination of isotope labeling with shotgun proteomics resulted in the total identification of 4434 protein clusters expressed in the microbial proteomic environment, 276 of which demonstrated differential abundance between control and IBD mice. Notably, application of a novel cysteine-reactive probe uncovered several microbial proteases and hydrolases overrepresented in the IBD mice. Implementation of these methods demonstrated that substantial insights into the identity and dysregulation of host and microbial proteins altered in IBD can be accomplished and can be used in the interrogation of other microbiome-related diseases

Quantitative Metaproteomics and Activity-Based Probe Enrichment Reveals Significant Alterations in Protein Expression from a Mouse Model of Inflammatory Bowel Disease

Tandem mass spectrometry based shotgun proteomics of distal gut microbiomes is exceedingly difficult due to the inherent complexity and taxonomic diversity of the samples. We introduce two new methodologies to improve metaproteomic studies of microbiome samples. These methods include the stable isotope labeling in mammals to permit protein quantitation across two mouse cohorts as well as the application of activity-based probes to enrich and analyze both host and microbial proteins with specific functionalities. We used these technologies to study the microbiota from the adoptive T cell transfer mouse model of inflammatory bowel disease (IBD) and compare these samples to an isogenic control, thereby limiting genetic and environmental variables that influence microbiome composition. The data generated highlight quantitative alterations in both host and microbial proteins due to intestinal inflammation and corroborates the observed phylogenetic changes in bacteria that accompany IBD in humans and mouse models. The combination of isotope labeling with shotgun proteomics resulted in the total identification of 4434 protein clusters expressed in the microbial proteomic environment, 276 of which demonstrated differential abundance between control and IBD mice. Notably, application of a novel cysteine-reactive probe uncovered several microbial proteases and hydrolases overrepresented in the IBD mice. Implementation of these methods demonstrated that substantial insights into the identity and dysregulation of host and microbial proteins altered in IBD can be accomplished and can be used in the interrogation of other microbiome-related diseases

Quantitative Metaproteomics and Activity-Based Probe Enrichment Reveals Significant Alterations in Protein Expression from a Mouse Model of Inflammatory Bowel Disease

Tandem mass spectrometry based shotgun proteomics of distal gut microbiomes is exceedingly difficult due to the inherent complexity and taxonomic diversity of the samples. We introduce two new methodologies to improve metaproteomic studies of microbiome samples. These methods include the stable isotope labeling in mammals to permit protein quantitation across two mouse cohorts as well as the application of activity-based probes to enrich and analyze both host and microbial proteins with specific functionalities. We used these technologies to study the microbiota from the adoptive T cell transfer mouse model of inflammatory bowel disease (IBD) and compare these samples to an isogenic control, thereby limiting genetic and environmental variables that influence microbiome composition. The data generated highlight quantitative alterations in both host and microbial proteins due to intestinal inflammation and corroborates the observed phylogenetic changes in bacteria that accompany IBD in humans and mouse models. The combination of isotope labeling with shotgun proteomics resulted in the total identification of 4434 protein clusters expressed in the microbial proteomic environment, 276 of which demonstrated differential abundance between control and IBD mice. Notably, application of a novel cysteine-reactive probe uncovered several microbial proteases and hydrolases overrepresented in the IBD mice. Implementation of these methods demonstrated that substantial insights into the identity and dysregulation of host and microbial proteins altered in IBD can be accomplished and can be used in the interrogation of other microbiome-related diseases

Triflic Acid Treatment Enables LC-MS/MS Analysis of Insoluble Bacterial Biomass

The lysis and extraction of soluble bacterial proteins from cells is a common practice for proteomics analyses, but insoluble bacterial biomasses are often left behind. Here, we show that with triflic acid treatment, the insoluble bacterial biomass of Gram^– and Gram⁺ bacteria can be rendered soluble. We use LC-MS/MS shotgun proteomics to show that bacterial proteins in the soluble and insoluble postlysis fractions differ significantly. Additionally, in the case of Gram^– Pseudomonas aeruginosa, triflic acid treatment enables the enrichment of cell-envelope-associated proteins. Finally, we apply triflic acid to a human microbiome sample to show that this treatment is robust and enables the identification of a new, complementary subset of proteins from a complex microbial mixture

Triflic Acid Treatment Enables LC-MS/MS Analysis of Insoluble Bacterial Biomass

The lysis and extraction of soluble bacterial proteins from cells is a common practice for proteomics analyses, but insoluble bacterial biomasses are often left behind. Here, we show that with triflic acid treatment, the insoluble bacterial biomass of Gram^– and Gram⁺ bacteria can be rendered soluble. We use LC-MS/MS shotgun proteomics to show that bacterial proteins in the soluble and insoluble postlysis fractions differ significantly. Additionally, in the case of Gram^– Pseudomonas aeruginosa, triflic acid treatment enables the enrichment of cell-envelope-associated proteins. Finally, we apply triflic acid to a human microbiome sample to show that this treatment is robust and enables the identification of a new, complementary subset of proteins from a complex microbial mixture

Identification of Regulatory Elements That Control PPARγ Expression in Adipocyte Progenitors

<div><p>Adipose tissue renewal and obesity-driven expansion of fat cell number are dependent on proliferation and differentiation of adipose progenitors that reside in the vasculature that develops in coordination with adipose depots. The transcriptional events that regulate commitment of progenitors to the adipose lineage are poorly understood. Because expression of the nuclear receptor PPARγ defines the adipose lineage, isolation of elements that control PPARγ expression in adipose precursors may lead to discovery of transcriptional regulators of early adipocyte determination. Here, we describe the identification and validation in transgenic mice of 5 highly conserved non-coding sequences from the PPARγ locus that can drive expression of a reporter gene in a manner that recapitulates the tissue-specific pattern of PPARγ expression. Surprisingly, these 5 elements appear to control PPARγ expression in adipocyte precursors that are associated with the vasculature of adipose depots, but not in mature adipocytes. Characterization of these five PPARγ regulatory sequences may enable isolation of the transcription factors that bind these <i>cis</i> elements and provide insight into the molecular regulation of adipose tissue expansion in normal and pathological states.</p></div

PPARγ CS1-5_<i>LacZ</i> positive cells express markers of adipose progenitors.

<p>Paraffin-embedded serial sections of X-gal stained subcutaneous WAT derived from PPARγ CS1-5_<i>LacZ</i> line 1 transgenic mice were analyzed by immunohistochemistry. Note that <i>LacZ</i> positive cells in transgenic fat pads express mural/endothelial/adipose progenitor cell markers (CD29, SMA), but not perilipin (mature adipocytes). Arrows point to several examples of the same <i>LacZ</i> positive cells in all serial sections, so that the overlap of markers can be evaluated.</p

The PPARγ CS1-5 cassette is transcriptionally active in white and brown fat cell progenitors.

<p>Paraffin-embedded sections of X-gal stained subcutaneous (<b>A,D,G</b>) and visceral (<b>B</b>,<b>E</b>,<b>H</b>) WAT, and BAT (<i>C,F,I</i>) from PPARγ (+/−),and PPARγ CS1-5_<i>LacZ</i> line 1 transgenic mice (6 weeks). Note the perivascular nature of many <i>LacZ</i> expressing cells in transgenic fat pads (arrows), and the strong blue stain in much of the vasculature of transgenic BAT (<b>F</b>). Genotypes indicated on top.</p