Long-read Proteogenomics Internal Standard triggered PRM (LRP-IS-PRM)
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Orbitrap Eclipse
ORGANISM(S): Homo Sapiens (human)
TISSUE(S): Stem Cell, Cell Culture
DISEASE(S): Disease Free
SUBMITTER: 
Erin Jeffery
LAB HEAD: Gloria Sheynkman
PROVIDER: PXD050909 | Pride | 2024-07-17
REPOSITORIES: Pride
Dataset's files
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Publications
IS-PRM-Based Peptide Targeting Informed by Long-Read Sequencing for Alternative Proteome Detection.
Korchak Jennifer A JA Jeffery Erin D ED Bandyopadhyay Saikat S Jordan Ben T BT Lehe Micah D MD Watts Emily F EF Fenix Aidan A Wilhelm Mathias M Sheynkman Gloria M GM
Journal of the American Society for Mass Spectrometry 20240716 11
Alternative splicing is a major contributor of transcriptomic complexity, but the extent to which transcript isoforms are translated into stable, functional protein isoforms is unclear. Furthermore, detection of relatively scarce isoform-specific peptides is challenging, with many protein isoforms remaining uncharted due to technical limitations. Recently, a family of advanced targeted MS strategies, termed internal standard parallel reaction monitoring (IS-PRM), have demonstrated multiplexed, s ...[more]
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Project description:Long-read proteogenomic data was used to create sample-matched protein database for WTC11 sample. This includes many potential alternative protein isoforms (major and minor isoforms) per gene. The IS-PRM method called Tomahto was tested versus DDA to demonstrate improved coverage of isoform-specific peptides. We called this overall method of long-read RNA informed Tomahto targeting LRP-IS-PRM and used it for protein-level evidence of multiple isoforms derived from alternative splicing.
2024-07-17 | PXD050904 | Pride
Project description:During the colonization of hosts, bacterial pathogens are presented with many challenges that must be overcome for colonization to successfully occur. This requires bacterial sensing of the surroundings and adaptation to the conditions encountered. One of the major impediments to pathogen colonization of the mammalian gastrointestinal tract is the antibacterial action of bile. Salmonella enterica serovar Typhimurium has specific mechanisms involved in resistance to bile. Besides being resistant to it, Salmonella can also successfully multiply in bile, using it as a source of nutrients. This accomplishment is highly relevant to pathogenesis, as Salmonella colonizes the gallbladder of hosts, where it can be carried asymptomatically and promote further host spread and transmission. In order to gain insights into the mechanisms used by Salmonella to grow in bile, we studied the changes elicited by Salmonella in the chemical composition of bile during growth in vitro and in vivo through a metabolomics approach. Our data suggest that phospholipids are an important source of carbon and energy for Salmonella during growth in the laboratory as well as during gallbladder infections of mice. Further studies in this area will generate a better understanding of how Salmonella exploits this generally hostile environment for its own benefit. For in vitro studies, bile was extracted from C57BL/6 mice and used immediately. An overnight culture of Salmonella enterica serovar Typhimurium SL1344 was used to inoculate 10 ?L of bile at an approximate density of 5x10^6 cells/mL. The experiment was performed in duplicate, and a total of 2 uninfected and 2 infected bile samples were studied. Samples were incubated for 24 hours at 37 oC with shaking. After incubation, samples were centrifuged to remove bacteria and the supernatant was saved for metabolomic analyses. For in vivo studies, C57BL/6 mice were infected with approximately 10^8 bacterial cells by oral gavage. Four groups of three to four mice each were either infected with Salmonella or kept uninfected (total of 11 mice per treatment group). Five days after infection, all mice were sacrificed and bile was collected. Samples were centrifuged and supernatants saved for metabolomic analyses. Samples were prepared by mixing equal volumes of bile from three to four mice, generating three samples per treatment (uninfected and infected), which were used in the subsequent steps. Seven microliters of each sample was evaporated and the residue resuspended in 50% acetonitrine. Extracts were infused into a 12-T Apex-Qe hybrid quadrupole-FT-ICR mass spectrometer equipped with an Apollo II electrospray ionization source, a quadrupole mass filter and a hexapole collision cell. Raw mass spectrometry data were processed as described elsewhere (Han et al. 2008. Metabolomics. 4:128-140). To identify differences in metabolite composition between different groups of samples, we filtered the list of masses for metabolites which were present on one set of samples but not the other. Additionally, we calculated the ratios between averaged intensities of metabolites from each group of mice. To assign possible metabolite identities to the masses selected as described above, the monoisotopic neutral masses of interest were queried against the Human Metabolome Database (HMDB, http://www.hmdb.ca), with a tolerance of 0.001 Da.
2011-07-06 | E-GEOD-30404 | biostudies-arrayexpress
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