Dataset Information

Novel Events, Gene Model Correction of Mycobacterium Avium by Proteogenomic Analysis

ABSTRACT: Mycobacterium avium is one of the prominent disease causing bacteria in humans. It causes lymphadenitis, chronic pulmonary and extrapulmonary and disseminated infections in adults, children and immunocompromised humans. M. avium has ~4,500 predicted gene models, out of which not all are identified at proteomic level. Proteomic database search followed by proteogenomic analysis helps in the correction of gene models, identification of novel exons/genes, variant proteins. As part of this study, we performed proteomic analysis of M. avium cultures by data-dependent acquisition (DDA) and data-independent acquisition (DIA) method followed by proteogenomic analysis of M. avium proteomic data. M. avium culture was subjected to proteomic sample preparation. The resulting peptides were acquired in 120min DDA (12 bRPLC) and DIA method using EASY nLC 1200 liquid chromatogram system coupled to Orbitrap Fusion Tribrid mass spectrometer. The resulting DDA raw files were searched sequentially against the M. avium proteome database, Mycobacterium tuberculosis H37Rv and Ra proteome database (Mtb), M. avium genome six-frame translated proteome and variant proteins database, respectively. The database search result was converted into a spectral library and used for DIA data search for the validation of GSSPs and variant peptides. The database search of M. avium DDA has resulted in the identification of 2,954 M. avium proteins, 128 Mtb proteins, 174 GSSPs (M. avium genome six-frame translated proteome) and 795 SNPs corresponding to 612 proteins (variant proteins database), respectively. The M. avium proteome database search has covered 62.09% of the proteome with the identification of 2,954 proteins out of 4,757 proteins in the database. From the manual categorization of 174 GSSPs, we observed 23 N-terminal extensions, 142 pseudogene coding peptides and one each novel exon and short peptide, respectively.

INSTRUMENT(S): Orbitrap Fusion

ORGANISM(S): Mycobacterium Avium 104

TISSUE(S): Cell Culture

SUBMITTER: Keshava Prasad T. S.

LAB HEAD: T. S. Keshava Prasad

PROVIDER: PXD032067 | Pride | 2023-03-17

REPOSITORIES: Pride

ACCESS DATA

Publications

Development of a Spectral Library for the Discovery of Altered Genomic Events in Mycobacterium avium Associated With Virulence Using Mass Spectrometry-Based Proteogenomic Analysis.

Kotimoole Chinmaya Narayana CN Antil Neelam N Kasaragod Sandeep S Behera Santosh Kumar SK Aravind Anjana A Reiling Norbert N Flo Trude Helen TH Prasad Thottethodi Subrahmanya Keshava TSK

Molecular & cellular proteomics : MCP 20230321 5

Mycobacterium avium is one of the prominent disease-causing bacteria in humans. It causes lymphadenitis, chronic and extrapulmonary, and disseminated infections in adults, children, and immunocompromised patients. M. avium has ∼4500 predicted protein-coding regions on average, which can help discover several variants at the proteome level. Many of them are potentially associated with virulence; thus, identifying such proteins can be a helpful feature in developing panel-based theranostics. In li ...[more]

PMID: 36948415

Similar Datasets

Project description:Background: Mycobacterium avium subspecies paratuberculosis (MPTb) is the causative agent of Johne’s disease, an intestinal disease of ruminants with major economic consequences. MPTb bacilli are phagocytosed by host macrophages upon exposure where they persist, resulting in lengthy subclinical phases of infection that can lead to immunopathology and disease dissemination. Consequently, analysis of the macrophage transcriptome in response to MPTb infection can provide valuable insights into the molecular mechanisms that underlie Johne’s disease. Here, we investigate pan-genomic gene expression in bovine monocyte-derived macrophages (MDM) purified from seven age-matched non-related females, in response to in vitro infection with MPTb (multiplicity of infection 2:1) at intervals of 2 hours, 6 hours and 24 hours post-infection. Results: Differentially expressed genes were identified by comparing the transcriptomes of the infected MDM to the control MDM at each time point (adjusted P-value threshold ≤ 0.10). 1,050 differentially expressed unique genes were identified 2 hours post-infection, with 974 and 84 differentially expressed unique genes detected 6 hours and 24 hours post-infection, respectively. Furthermore, in the infected MDM the number of upregulated genes exceeded the number of downregulated genes at each time point, with the fold-change in expression for the upregulated genes markedly higher than that for the downregulated genes. Conclusions: Inspection and systems analysis of the differentially expressed genes revealed changes in the expression profiles of genes involved in the inflammatory response, cell signalling pathways and apoptosis. The transcriptional changes associated with cellular signalling and the inflammatory response may reflect different immuno-modulatory mechanisms that underlie host-pathogen interactions during infection. Affymetrix GeneChip® Bovine Genome Arrays were used to examine gene expression of bovine monocyte-derived macrophages (MDM) after in vitro challenge with Mycobacterium avium subspecies paratuberculosis across a time series of 2 hr, 6 hr and 24 hr post-challenge. A 0 hr control treatment was also generated and seven different age-matched female Holstein-Friesian cattle were used for each time-point/treatment combination for a total of 49 microarrays, one of which was excluded from analysis since it did not pass quality control.

Project description:Investigation of whole genome gene expression level changes in BMDM adhered for 4 or 18hrs to Lab Tek Chamber slides pre-coated with 4ug/ml of human serum albumin (HSA) (control protein) or purified human C1q and treated with lipopolysaccharide (LPS: 20ng/ml) Mycobacterium avium 101 (M avium) or apoptotic Jurkat T cells. BMDM were obtained from C57Bl/6 and generated as previously described in Bohlson, SS, Strasser, JA, Bower JJ, Schorey J S 2001 Role of complement in Mycobacterium avium pathogenesis: in vivo and in vitro analyses of the host response to infection in the absence of complement component C3 Infec Immun 69: 7729-7735. Mybacterium avium 101 was obtained from Dr Jeff Schorey (University of Notre Dame). The human Jurkat T cell line was obtained from ATCC (Manassas, VA) and induced to undergo apoptosis with dexamethasone as described in Lillis, AP, Greenlee, M C, Mikhailenko I, Pizzo, S V, Tenner, A J, Strickland, D K, Bohlson, S S 2008 Murine Low-Density Lipoprotein Receptor-Related Protein 1 (LRP) Is Required for Phagocytosis of Targets Bearing LRP Ligands but Is Not Required for C1q-triggered Enhancement of Phagocytosis J Immunol 181: 364: 373. A six chip study using total RNA from 5 separate cultures of BMDM adhered to HSA for 4 hrs, 5 separate cultures of BMDM adhered to C1q for 4 hrs, 5 separate cultures of BMDM adhered to HSA for 4 hrs and infected with M avium at a 1:500 ratio of BMDM to M avium, 5 separate cultures of BMDM adhered to C1q for 4hrs and infected with M avium at a 1:500 ratio ofBMDM to M avium, 5 separate cultures of BMDM adhered to HSA for 4 hrs and infected with M avium at a 1:1000 ratio of BMDM to M avium, 5 separate cultures of BMDM adhered to HSA for 4 hrs and treated with 20ng/ml LPS, 5 separate cultures of BMDM adhered to C1q for 4 hrs and treated with 20ng/ml LPS, 5 separate cultures of BMDM adhered to HSA for 4 hrs and treated with 20ng/ml LPS and co-cultured with apoptotic Jurkat cells at a 1:3 ratio of BMDM to apoptotic cells, 5 separate cultures of BMDM adhered to C1q for 4 hrs and treated with 20ng/ml LPS and co-cultured with apoptotic Jurkat cells at a 1:3 ratio of BMDM to apoptotic cells, 5 separate cultures of BMDM adhered to HSA for 18 hrs, 5 separate cultures of BMDM adhered to C1q for 18 hrs, 5 separate cultures of BMDM adhered to HSA for 18 hrs and infected with M avium at a 1:500 ratio of BMDM to M avium, 5 separate cultures of BMDM adhered to C1q for 18hrs and infected with M avium at a 1:500 ratio of BMDM to M avium, 5 separate cultures of BMDM adhered to HSA for 18 hrs and infected with M avium at a 1:1000 ratio of BMDM to M avium. The chip utilized for these studies is a mouse whole-genome 12-plex expression microarray design by NimbleGen designed from the MM9 genome Candidate probe sequences were verified to have no cross-hybridization to human (HG19) or Mycobacterium avium (NC_008595) targets Note: Study sample BMDM_HSA & M avium (1:500)_18hr_rep2 was not included in this submission due to quality control concerns.

Dataset Information

Novel Events, Gene Model Correction of Mycobacterium Avium by Proteogenomic Analysis

Publications

Development of a Spectral Library for the Discovery of Altered Genomic Events in Mycobacterium avium Associated With Virulence Using Mass Spectrometry-Based Proteogenomic Analysis.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure