Decoding the genomic basis of osteoarthritis
Ontology highlight
ABSTRACT:
INSTRUMENT(S): LTQ Orbitrap
ORGANISM(S): Homo Sapiens (human)
SUBMITTER: 
James Wright
LAB HEAD: Jyoti Choudhary
PROVIDER: PXD014666 | Pride | 2020-11-17
REPOSITORIES: Pride
Similar Datasets
Project description:The aim of this work is to apply an integrated systems approach to understand the biological underpinnings of hip osteoarthritis that culminates in the need for total joint replacement (TJR). This study is a feasibility pilot that integrates functional genomics data from diseased and non-diseased tissues of OA patients who have undergone TJR. For each tissue, we characterised epigenetic marks (methylation), gene transcription (RNASeq) and expression (quantitative proteomics). We also generated genotype data on the HumanCoreExome array for each individual. This data is part of a pre-publication release.
2018-05-14 | PXD006673 | Pride
Project description:Large scale proteomic profiling of cell lines can yield valuable insights into the molecular signatures attributed to variable genotypes or induced perturbations. Specifically, the ability to perform deep and rapid proteome analysis of pharmacologically modulated cells could generate drug-protein associations for large libraries of compounds that predict mechanism of action and enable rational drug design. Although isobaric labelling has greatly increased the throughput of proteomic analysis at deep coverage, the commonly used sample preparation workflows often require complex time-consuming steps and/or costly consumables, limiting their suitability for large scale studies. Here, we present a simplified and cost effective one-pot reaction sample preparation workflow in a 96-well plate format with manual parallel processing (SimPLIT), that minimizes processing steps and reduces technical variability. The workflow is based on a sodium deoxycholate lysis buffer and a single detergent clean-up step after peptide labeling, followed by quick off-line fractionation and MS2 analysis. The simplified workflow demonstrates high reproducibility and provides improved proteome representation compared to alternative approaches. We showcase the large-scale applicability of the workflow by investigating proteomic heterogeneity in a panel of colorectal cancer cell lines and by performing target discovery for a set of molecular glue degraders in different cell lines, in a 96-sample assay. Using this workflow, we report a subset of frequently dysregulated proteins in colorectal cancer cells and uncover cell-dependent protein degradation profiles of seven cereblon E3 ligase modulators (CRL4CRBN). Overall, SimPLIT is a robust method that can be easily implemented in most proteomics laboratories for medium-to-large scale TMT-based studies involving deep profiling of cell lines.
2022-08-03 | PXD034022 | Pride
Project description:Large scale proteomic profiling of cell lines can yield valuable insights into the molecular signatures attributed to variable genotypes or induced perturbations. Specifically, the ability to perform deep and rapid proteome analysis of pharmacologically modulated cells could generate drug-protein associations for large libraries of compounds that predict mechanism of action and enable rational drug design. Although isobaric labelling has greatly increased the throughput of proteomic analysis at deep coverage, the commonly used sample preparation workflows often require complex time-consuming steps and/or costly consumables, limiting their suitability for large scale studies. Here, we present a simplified and cost effective one-pot reaction sample preparation workflow in a 96-well plate format with manual parallel processing (SimPLIT), that minimizes processing steps and reduces technical variability. The workflow is based on a sodium deoxycholate lysis buffer and a single detergent clean-up step after peptide labeling, followed by quick off-line fractionation and MS2 analysis. The simplified workflow demonstrates high reproducibility and provides improved proteome representation compared to alternative approaches. We showcase the large-scale applicability of the workflow by investigating proteomic heterogeneity in a panel of colorectal cancer cell lines and by performing target discovery for a set of molecular glue degraders in different cell lines, in a 96-sample assay. Using this workflow, we report a subset of frequently dysregulated proteins in colorectal cancer cells and uncover cell-dependent protein degradation profiles of seven cereblon E3 ligase modulators (CRL4CRBN). Overall, SimPLIT is a robust method that can be easily implemented in most proteomics laboratories for medium-to-large scale TMT-based studies involving deep profiling of cell lines.
2022-08-03 | PXD031510 | Pride
Project description:Assessing the impact of genomic alterations on protein networks is fundamental in identifying the mechanisms that shape cancer heterogeneity. We have used isobaric labelling to characterize the proteomic landscapes of 50 colorectal cancer cell lines and to decipher the functional consequences of somatic genomic variants. The robust quantification of over 9,000 proteins and 11,000 phosphopeptides on average, enabled the de novo construction of a functional protein correlation network which ultimately exposed the collateral effects of mutations on protein complexes. CRISPR-cas9 deletion of key chromatin modifiers confirmed that the consequences of genomic alterations can propagate through protein interactions in a transcript-independent manner. Lastly, we leveraged the quantified proteome to perform unsupervised classification of the cell lines and to build predictive models of drug response in colorectal cancer. Overall, we provide a deep integrative view of the functional network and the molecular structure underlying the heterogeneity of colorectal cancer cells.
2017-08-09 | PXD005235 | Pride
Project description:Parkinson's disease (PD) is a growing burden worldwide, and despite ongoing efforts to find reliable biomarkers for early and differential diagnosis, prognosis and disease monitoring, there is no biofluid biomarker used in clinical routine to date. Cerebrospinal fluid (CSF) is collected often and should closely reflect structural and functional alterations in PD patients' brains. Here we describe a scalable and sensitive mass spectrometry (MS)-based proteomics workflow for CSF proteome profiling to find specific biomarkers and identify disease-related changes in CSF protein levels in PD. From two independent cohorts consisting of more than 200 individuals, our workflow reproducibly quantified over 1,700 proteins from minimal sample amounts. Combined with machine learning, this identified a group of several proteins, including OMD, CD44, VGF, PRL, and MAN2B1 that were altered in PD patients or significantly correlate with clinical scores, indicative of disease progression. Interestingly, we uncovered signatures of enhanced neuroinflammation in patients with familial PD (LRRK2 G2019S carriers) as indicated by increased levels of CTSS, PLD4, HLA-DRA, HLA-DRB1, and HLA-DPA1. A comparison with urinary proteome changes in PD patients revealed a large overlap in protein composition PD-associated changes in these body fluids, including lysosomal factors like CTSS. Our results validate MS-based proteomics of CSF as a valuable strategy for biomarker discovery and patient stratification in a neurodegenerative disease like PD. Consistent proteomic signatures across two independent CSF cohorts and previously acquired urinary proteome profiles open up new avenues to improve our understanding of PD pathogenesis.
2022-06-01 | PXD026491 | Pride
Project description:The mechanisms by which the mouse pathogen Citrobacter rodentium triggers effacement of the brush border microvilli, colitis, hyperplasia and dysbiosis remain poorly understood. We investigated the impact of C. rodentium infection on the proteomic and metabolic landscapes of intestinal epithelial cells (IECs) in vivo using isobaric labeling proteomics and targeted metabolomics. We found that infection depletes proteins involved in butyrate uptake, glycolysis, TCA cycle, lipid metabolism and oxidative phosphorylation with aparallel, increased production of creatine/phosphocreatine and cholesterol. The evolving ecological niche within the infected gut was concomitant with a reduction in butyrate-producing commensal bacteria and expansion of Proteobacteria that can metabolize cholesterol. These changes coincide with the modulation of IEC transcription factors by C. rodentium, specifically phosphorylation of Kdm5a, demethylation of histone H3 (Lys-4) and cleavage of Srebp2. Taken together our results show that whilst engaging with the host in a race to control innate immune responses, C. rodentium and the changing microbiota shape the metabolism flow in IECs.
2018-01-10 | PXD005004 | Pride
Project description:We used the mouse pathogen Citrobacter rodentium to model gut infections. Following oral inoculation C. rodentium resides in the caecum for the first 3 days, before it infects the colon on the 4th day. Here we show that while the host is unresponsive to the infection on day 3, there is an abrupt reprogramming of the cellular composition of the crypt, involving depletion of goblet and deep crypt secretory cells, as well as metabolism (e.g. simultaneous up-regulation of cholesterol biogenesis, import and efflux), DNA damage repair and proliferation, which correlated with Ki67 staining, on day 4. Reduction in the abundance of proteins involved in the TCA cycle and oxidative phosphorylation, leading to oxygenation of the gut, coincided with instant expansion of mucosal-associated Enterobacteriaceae. These results show that sensing a small number of pathogenic bacteria triggers immediate intrinsic changes to the epithelium physiology and the microbiota, which parallel innate gut immune responses.
2019-03-05 | PXD012031 | Pride
Project description:Background: In autosomal dominant polycystic kidney disease (ADPKD) there is an unmet need for early markers of rapid disease progression to facilitate counseling and selection for kidney-protective therapy. Our aim was to identify markers for rapid disease progression in urinary extracellular vesicles (uEVs). Methods: uEVs were isolated at baseline in two independent cohorts including patients with rapid disease progression and stable disease; patients were matched by age, sex, total kidney volume and mutation. uEV biomarker candidates were identified by mass spectrometry and further analyzed in a third and fourth cohort using immunoblotting and an enzyme-linked immunosorbent assay (ELISA). Single-nucleotide RNA sequencing of healthy and ADPKD tissue was used to confirm findings and identify the cellular origin of the uEV biomarker. Results: In the first two cohorts matrix metalloproteinase-7 (MMP-7) was significantly higher in uEVs of patients with rapid disease progression compared to stable disease. In the third cohort, immunoblotting confirmed a >2-fold increase in uEV-MMP-7 in patients with rapid disease progression compared to stable disease. In the fourth cohort, whole urine rather than uEVs were analyzed with MMP-7 ELISA demonstrating a significant but weak correlation with kidney function decline. Single-nucleotide RNA sequencing showed higher MMP-7 expression in ADPKD kidney tissue than in healthy kidney tissue and localized MMP-7 to the proximal tubule and thick ascending limb. Conclusion: uEV-associated MMP-7 is elevated in ADPKD patients with rapid disease progression and should be further investigated as predictive biomarker. Our findings also suggests that MMP-7 performs better when analyzed in uEVs than in whole urine.
2023-12-18 | PXD042533 | Pride
Project description:Aneuploidy results in decreased cellular fitness in many different species and model systems. However, aneuploidy is commonly found in cancer cells and often correlates with aggressive growth and poor prognosis, suggesting that the impact of aneuploidy on cellular fitness is context dependent. The BRG1 (SMARCA4) subunit of the SWI/SNF chromatin remodelling complex is a tumour suppressor that is frequently lost in cancer cells. Here, we used a chromosomally stable cell line to test the effect of BRG1 loss on the evolution of aneuploidy. We find that BRG1 deletion leads to an initial loss of fitness in this cell line that improves over time. The improved fitness correlates with a gain of extra copies of chromosome 18. Notably, changes in pathways that are known to promote tolerance to aneuploidy are evident immediately upon loss of BRG1, providing an environment where karyotype changes associated with a fitness advantage can be explored. At least in some genetic backgrounds, therefore, loss of the SWI/SNF complex can contribute to tumourigenesis through tolerance of aneuploidy.
2022-03-23 | PXD014954 | Pride
Project description:Citrobacter rodentium is commonly used to elucidate mucosal responses to infection in mice developing mild disease (e.g. C57BL/6), while little is known about host responses to infection in mice developing severe disease (e.g. C3H/HeN). We report that the phyla Bacteroidetes is a minor component of the tissue-associated microbiome in uninfected C3H/HeN mice. Following infection, C. rodentium rapidly and uniformly colonises the C3H/HeN colonic mucosa, which coincides with downregulation of proteins involved in the TCA cycle and oxidative phosphorylation in intestinal epithelial cells (IECs). In contrast, we observed upregulation of DNA replication and DNA damage repair processes, as well as cholesterol biogenesis, import and export, nutritional immunity, IL-22 and INFg responses, and expression of NLRP3, in IECs. Moreover, C. rodentium triggers a staggered cell proliferation response from 3 days post infection, which correlated with a higher abundance of SLC5A9 and reduced abundance of the IEC differentiation markers SLC26A3 and CA4. Uniquely, C. rodentium triggers differential secretion of gel-forming mucins, with the number of goblet cells filled with acidic and neutral mucins dramatically increasing and decreasing, respectively. Together, these results show that despite vigorous responses, C3H/HeN mice succumb to C. rodentium infection, possibly as a result of excessive and disordered mucosal responses.
2020-01-21 | PXD012846 | Pride