ABSTRACT: Proteomic expression analysis of Ca. Lokiarchaeum ossiferum, strain B35, grown using casein hydrolysate as the main carbon source under anaerobic conditions
Project description:Proteome characterization of three E. coli strains: a methanol-dependent strain (MEvo1-0) and two synthetically methylotrophic strains (MEcoli_ref_1, MEcoli_ref_2)
Project description:Using a combination of proteomics, interaction studies and structural characterization we show that WDR73 and BRAT1 are interactors of the heterodimer INTS9-11 of INT. Both proteins are involved in the stability and formation of the cleavage module of INT consisting of INTS4-9-11.
Project description:Background. Dyslipidemia is associated with chronic kidney disease (CKD). It has been shown that inhibition of β-oxidation and lipid accumulation are common threats in the onset of kidney injury and conducive to fibrosis. We characterized the S-acylated proteome of kidneys from mice with diet-induced nephropathy, to uncover novel lipid-related biochemical changes that might contribute to the lipid-induced kidney dysfunction. Methods. We conducted total and S-acylated proteomics, and lipidomics of the renal crude membrane fractions of mice fed a high-fat diet (HFD). Then, we investigated in vitro the effect of S-acylation inhibition on megalin expression and ligand binding. Results. HFD caused albuminuria with unchanged kidney function. The dietary intervention induced a large-scale repression of protein S-acylation as well as of the most abundant ceramides and sphingomyelin species, which are highly suggestive of an overall reduction in acyl-CoA availability. S-acylation repression mostly affected proteins involved in endocytosis and intracellular transport. In particular, the kidney of the mice fed a HFD, which are characterized by albuminuria, displayed a marked decrease in the total amount and in the S-acylated form of megalin, the main tubular protein retrieval system. Further in vitro experiments indicated that S-acylation is important for total expression of megalin. Conclusions. The diet-induced derangement of fatty acids metabolism modifies the renal landscape of the S-acylated proteome during early stage of the kidney injury, which might reduce the maximal capacity of protein reabsorption by the proximal tubule.
Project description:Aims/hypothesis AGEs are considered environmental contributors of type 1 diabetes, but the exact role AGEs play early in pathogenesis of the disease, remains unidentified. We aimed to reduce AGEs with the pharmacotherapy alagebrium chloride in MIN6N8 cells and early in life in NOD mice to determine its impact on beta cell immunogenicity, function, and disease progression. Methods MIN6N8 cells were cultured with AGEs with or without short-term alagebrium therapy to determine the effect on ER stress and beta cell antigen presentation via a reporter assay for protein responses in the unfolded protein response, the enzymatic activity of endoplasmic reticulum aminopeptidase-1 (ERAP1) and the immunopeptidome. To determine the effect of short-term alagebrium therapy on beta cells in vivo, female NOD mice were treated with alagebrium and insulin secretion, insulitis, and immune cell repertoire were studied. Adoptive transfer studies and diabetes progression studies were used to consider the impact of alagebrium on immune cell function and disease outcome. Results In MIN6N8 cells, alagebrium therapy inhibited the induction of endoplasmic reticulum (ER) stress and the activity of ERAP1 by AGE-modified proteins. Alagebrium treated-MIN6N8 cells did not change the MHC Class I presentation of known beta cell antigens but did induce proteomic changes related to ER homeostatic pathways. Prior to overt autoimmune diabetes, female NOD mice treated with alagebrium for 30-40 days had improved insulin secretion, reduced insulitis and amplified proportions of pancreatic CD8+ T cells, mature B cells and F4/80+ macrophages. Splenocytes from alagebrium-treated mice adoptively transferred disease to NODscid recipients and maintained interferon production in vitro. While partial protection of islets by alagebrium was seen following the adoptive transfer of activated NOD G9C8 transgenic TCR CD8+ T cells, alagebrium therapy in NOD mice did not reduce diabetes progression. Conclusions/interpretation Our data suggests that in experimental models of diabetes, short-term alagebrium treatment allows for beta cell improvement, and maintenance of immune cell function, which does not fully mitigate diabetes progression.
Project description:We have analysed a patinet derived mesothelioma cell line by immuniopeptidomics and identified cancer antignes in this line immunopeptidome.
Project description:Peptide vaccination remains a viable approach to induce T-cell mediated killing of tumours. To identify potential T-cell targets for Triple-Negative Breast Cancer (TNBC) vaccination, we examined the effect of the pro-inflammatory cytokine interferon-γ (IFNγ) on the transcriptome, proteome and immunopeptidome of the TNBC cell line MDA-MB-231. Using high resolution mass spectrometry, we identified a total of 84,131 peptides from 9,647 source proteins presented by human leukocyte antigen (HLA)-I and HLA-II alleles. Treatment with IFNγ resulted in a remarkable remoulding of the immunopeptidome, with only a 34% overlap between untreated and treated cells across the HLA-I immunopeptidome, and expression of HLA-II only on treated cells. IFNγ increased the overall number, diversity and abundance of the immunopeptidome, as well as the proportion of coverage of source antigens. The suite of peptides displayed under conditions of IFNγ treatment included many known tumour associated antigens, with the HLA-II repertoire sampling 265 breast cancer associated antigens absent from those sampled by HLA-I. Quantitative analysis of the transcriptome (10,248 transcripts) and proteome (6783 proteins) of these cells revealed 229 proteins and transcripts were commonly differentially expressed, most of which involved in downstream targets of IFNγ signalling including components of the antigen processing machinery such as tapasin and HLA. However, these changes in protein expression did not explain the dramatic modulation of the immunopeptidome following IFNγ treatment. These results demonstrate the high degree of plasticity in the immunopeptidome TNBC cells following cytokine stimulation and provide evidence that under pro-inflammatory conditions a greater variety of HLA-I and HLA-II vaccine targets are unveiled to the immune system. This has important implications for the development of personalised cancer vaccination strategies.
Project description:Abstract Background Vertebral endplate signal intensity changes visualized by magnetic resonance imaging termed Modic changes (MC) are highly prevalent in low back pain patients. Interconvertibility between the three MC subtypes (MC1, MC2, MC3) suggests different pathological stages. Histologically, granulation tissue, fibrosis, and bone marrow edema are signs of inflammation in MC1 and MC2. However, different inflammatory infiltrates and amount of fatty marrow suggest distinct inflammatory processes in MC2. Aims The aims of this study were to investigate i) the degree of bony (BEP) and cartilage endplate (CEP) degeneration in MC2, ii) to identify inflammatory MC2 pathomechanisms, and iii) to show that these marrow changes correlate with severity of endplate degeneration. Methods Pairs of axial biopsies (n=58) spanning the entire vertebral body including both CEPs were collected from human cadaveric vertebrae with MC2. From one biopsy, the bone marrow directly adjacent to the CEP was analyzed with mass spectrometry. Differentially expressed proteins (DEPs) between MC2 were identified and bioinformatic enrichment analysis was performed. The other biopsy was processed for paraffin histology and BEP/CEP degenerations were scored. Endplate scores were correlated with DEPs. Results Endplates from MC2 were significantly more degenerated. Proteomic analysis revealed an activated complement system, increased expression of extracellular matrix proteins, angiogenic, and neurogenic factors in MC2 marrow. Endplate scores correlated with upregulated complement and neurogenic proteins. Discussion The inflammatory pathomechanisms in MC2 comprises activation of the complement system. Concurrent inflammation, fibrosis, angiogenesis, and neurogenesis indicate that MC2 is a chronic inflammation. Correlation of endplate damage with complement and neurogenic proteins suggest that complement system activation and neoinnervation may be linked to endplate damage. The endplate-near marrow is the pathomechanistic site, because MC2 occur at locations with more endplate degeneration. Conclusion MC2 are fibroinflammatory changes with complement system involvement which occur adjacent to damaged endplates.
Project description:Tripartite Tc toxins are virulence factors of bacterial pathogens. Although their structure and mechanism of action are well understood, it remains elusive where this large macromolecular complex is assembled and how it is released. Here we show by an integrative multiscale imaging approach that Yersinia entomophaga Tc (YenTc) toxin components are expressed only in a subpopulation of cells that are “primed” with several other potential virulence factors, including filaments of the protease M66/StcE. A phage-like lysis cassette (LC) is required for YenTc release; however, before resulting in complete cell lysis, the LC generates intermediate “ghost” cells, which may serve as assembly compartments and become densely packed with assembled YenTc holotoxins. We hypothesize that this stepwise mechanism evolved to minimize the number of cells that need to be sacrificed. The occurrence of similar lysis cassettes in diverse organisms indicates a conserved mechanism for Tc toxin release that may apply to other extracellular macromolecular machines.
Project description:MTCH2 is a protein localized in the outer membrane of mitochondria. It belongs to the solute carrier 25 family, but its substrates or transporter function remains unknown. Previous research links MTCH2 to apoptosis induction, and Alzheimer’s disease, mitochondrial metabolism and dynamics, and MTCH2 has been shown to function as a protein insertase and scramblase. Moreover, MTCH2 is a significant regulator of adipocyte differentiation and lipid homeostasis. Genome-wide association studies have identified MTCH2 variants to be associated with increased body mass index, obesity, and diabetes risk. Thus, MTCH2 emerges as a promising candidate for modulating adipocyte function and whole-body energy metabolism, but its specific metabolic role in mature adipose tissues remains unexplored. In this study, we show that MTCH2 regulates mitochondrial function and whole-body energy expenditure by regulating lipid utilization in adipose tissue.