Mass Spectrometry Footprinting Reveals How Kinetic Stabilizers Counteract Transthyretin Dynamics Altered by Pathogenic Mutations_FPOP
Ontology highlight
ABSTRACT:
INSTRUMENT(S):
ORGANISM(S): Homo Sapiens (human)
SUBMITTER: 
RAVI KANT
LAB HEAD: Dr. Michael L. Gross
PROVIDER: PXD071928 | Pride | 2025-12-13
REPOSITORIES: Pride
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| Processing_Outputs.zip | Other | |||
| Raw_Data.zip | Other | |||
| V122I.fasta | Fasta | |||
| V30M.fasta | Fasta | |||
| WT.fasta | Fasta |
Items per page: 5 1 - 5 of 6 |
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Project description:Mass Spectrometry Footprinting Reveals How Kinetic Stabilizers Counteract Transthyretin Dynamics Altered by Pathogenic Mutations
2025-12-13 | PXD071902 | Pride
Project description:Age-related wild type transthyretin amyloidosis (wtATTR) is characterized by systemic deposition of amyloidogenic fibrils of misfolded transthyretin (TTR) in the connective tissue of many organs. In the heart this leads to cardiac dysfunction, which is a significant cause of age-related heart failure. The hypothesis tested is that TTR affects cardiac fibroblasts in ways that may contribute to fibrosis. When primary cardiac fibroblasts were cultured on TTR-deposited substrates, the F-actin cytoskeleton disorganized, focal adhesion formation decreased, and nuclear shape was flattened. Fibroblasts had faster collective and single cell migration velocities on TTR-deposited substrates. Additionally, fibroblasts cultured on microposts with TTR deposition had reduced attachment and increased proliferation above untreated. Transcriptomic and proteomic analyses of fibroblasts grown on glass covered with TTR showed significant upregulation of inflammatory genes after 48 hours, indicative of progression in TTR-based diseases. Together, results suggest that TTR deposited in tissue extracellular matrix may affect both the structure, function and gene expression of cardiac fibroblasts. As therapies for wtATTR are cost-prohibitive and only slow disease progression, better understanding of cellular maladaptation may elucidate novel therapeutic targets.
2021-06-02 | GSE175917 | GEO
Project description:The biological effects of TTR proteins in the vasculature remain unknown. We used microarrays to detail the modulation of gene expression on HUVECs by V30M TTR when compared to cells exposed to WT TTR. HUVECs (passage 7) were cultured in the presence of WT or V30M TTR at 4M-BM-5M for 3 hours. RNA was extracted and hybridized on Affymetrix microarrays. We sought to obtain differentially expressed genes modulated by V30M TTR protein.
2013-09-13 | E-GEOD-44856 | biostudies-arrayexpress
Project description:Transthyretin amyloid cardiomyopathy (ATTR-CM) is characterized by the misfolding of transthyretin (TTR), fibrillogenesis, and progressive amyloid fibril deposition in the myocardium, leading to cardiac dysfunction with dismal prognosis. In ATTR-CM, either destabilizing mutations (variant TTR, ATTRv) or ageing-associated processes (wild-type TTR, ATTRwt) lead to the formation of TTR amyloid fibrils. Due to a lack of representative disease models, ATTR-CM disease mechanisms are largely unknown, thereby limiting disease understanding and therapeutic discovery. Methods and Results: Here, we report a novel in vitro ATTR-CM model which uncovers cell type-specific disease phenotypes by exposing the three major human cardiac cell types to TTR fibrils, thereby providing novel insights into the cellular mechanisms of ATTR-CM disease. Human recombinant TTR proteins (WT, V122I, V30M) and respective fibrils were generated and characterized using Thioflavin T, Amytracker, Congo red and dot blot analyses. Seeding human induced pluripotent stem cell-derived-cardiomyocytes (hiPSC-CMs) and endothelial cells (ECs) on TTR fibrils resulted in reduced cell viability. Confocal microscopy revealed extracellular localization of TTR fibrils to hiPSC-CMs, leading to sarcomere disruption, altered calcium handling and disrupted electromechanical coupling, while ECs showed a reduced migration capacity with aberrant cell morphology. hiPSC-fibroblasts (hiPSC-FBs) were largely unaffected by TTR fibrils, presenting normal viability, but showing enhanced localization with TTR fibrils. Our model shows that WT and variant TTR fibrils lead to cell type-specific phenotypes, providing novel insights into the underlying cellular disease mechanisms of ATTR-CM, thereby facilitating the identification of novel therapeutic targets and biomarkers.
2025-06-11 | GSE294021 | GEO
Project description:Early diagnosis of transthyretin (TTR) amyloid diseases remains challenging because of variable disease penetrance. Currently, patients must have an amyloid positive tissue biopsy to be eligible for disease modifying therapies. Early diagnosis is often difficult because the patient exhibits apparent symptoms of polyneuropathy or cardiomyopathy, but has a negative amyloid biopsy. Thus, there is a pressing need for more objective, quantitative diagnostics and biomarkers of TTR-aggregation-associated polyneuropathy and cardiomyopathy. This is especially true in the context of clinical trials demonstrating significant disease modifying effects, e.g. when the TTR tetramer stabilizer tafamidis was administered to familial amyloid polyneuropathy (FAP) patients early in the disease course. When asked if the findings of the tafamidis registration trial were “sufficiently robust to provide substantial evidence of efficacy for a surrogate endpoint that is reasonably likely to predict a clinical benefit” the advisory committee said yes, but the FDA rejected the tetramer stabilization surrogate biomarker required for orphan tafamidis approval–hence, acceptable biomarkers are badly needed. Herein, we explored whether peripheral blood cell mRNA expression profiles could differentiate symptomatic from asymptomatic V30M FAP patients, and if such a profile would normalize upon tafamidis treatment. We demonstrate that blood cell gene expression patterns reveal sex-independent as well as male and female specific inflammatory signatures in symptomatic FAP patients, but not in asymptomatic carriers, that normalize in FAP patients 6 months after tafamidis treatment. Thus these signatures have potential both as an early diagnostic and as a surrogate biomarker for measuring response to treatment in FAP patients.
2016-08-09 | GSE67784 | GEO
Project description:Early diagnosis of transthyretin (TTR) amyloid diseases remains challenging because of variable disease penetrance. Currently, patients must have an amyloid positive tissue biopsy to be eligible for disease modifying therapies. Early diagnosis is often difficult because the patient exhibits apparent symptoms of polyneuropathy or cardiomyopathy, but has a negative amyloid biopsy. Thus, there is a pressing need for more objective, quantitative diagnostics and biomarkers of TTR-aggregation-associated polyneuropathy and cardiomyopathy. This is especially true in the context of clinical trials demonstrating significant disease modifying effects, e.g. when the TTR tetramer stabilizer tafamidis was administered to familial amyloid polyneuropathy (FAP) patients early in the disease course. When asked if the findings of the tafamidis registration trial were âsufficiently robust to provide substantial evidence of efficacy for a surrogate endpoint that is reasonably likely to predict a clinical benefitâ the advisory committee said yes, but the FDA rejected the tetramer stabilization surrogate biomarker required for orphan tafamidis approvalâhence, acceptable biomarkers are badly needed. Herein, we explored whether peripheral blood cell mRNA expression profiles could differentiate symptomatic from asymptomatic V30M FAP patients, and if such a profile would normalize upon tafamidis treatment. We demonstrate that blood cell gene expression patterns reveal sex-independent as well as male and female specific inflammatory signatures in symptomatic FAP patients, but not in asymptomatic carriers, that normalize in FAP patients 6 months after tafamidis treatment. Thus these signatures have potential both as an early diagnostic and as a surrogate biomarker for measuring response to treatment in FAP patients. Study Subjects: Tafamidis-treated and untreated V30M FAP patients, asymptomatic V30M carriers, and healthy, age- and sex-matched controls without TTR mutations had whole blood drawn. RNA was extracted from a total of 309 peripheral blood samples collected in PAXgene tubes (Qiagen). RNA Extraction and Microarray Processing: RNA was extracted with the PAXgene RNA Whole Blood Kit. Each RNA sample (100ng) was amplified and labeled with the Ambion WT Express Kit and hybridized to Affymetrix HuGene 1.1 ST Arrays using the Affymetrix GeneTitan Multi-Channel (MC) platform. Normalized signals were generated using Robust Multichip Average (RMA) in Partek Genomics Suite software version 6.6. Probesets on the DNA microarrays with low expressed signals were determined using a kernel density plot, and signals <Log2 of 6 were excluded from analysis.
2016-08-09 | E-GEOD-67784 | biostudies-arrayexpress
Project description:We have developed a new mouse model of transthyretin (TTR) amyloidosis using transgenic mice expressing the most fibrillogenic variant of TTR (S52P). Following seeding with amyloid fibrils, TTR amyloid is deposited mainly in the heart and tongue. The fibrils contain both full length and truncated (49-127) TTR. The presence of S52P TTR was confirmed by proteomics. Knockout of alpha2-antiplasmin enhanced amyloid deposition.
2021-12-27 | PXD027747 | Pride
Project description:Background: Chronic stress significantly contributes to mood- and anxiety disorders. Previous data suggest a correlative connection between vitamin B12 supplementation, depression, and stress resilience. However, the underlying mechanisms are still poorly understood. Methods: Using the chronic variable stress mouse model coupled with RNA-sequencing, we determined vitamin B12-induced transcriptional changes related to stress resilience. By viral-mediated gene transfer and in vivo epigenome editing, we reveal a functional pathway linking vitamin B12, DNA methylation, and depressive-like symptoms. Results: We identified Transthyretin (Ttr) as a sex-specific key target of vitamin B12 action in chronic stress. Accordingly, TTR expression was increased postmortem in the prefrontal cortex of male, but not female, depressed patients. Virally altered Ttr in the prefrontal cortex functionally contributed to stress- and depression-related behaviors, changes in dendritic spine morphology, and gene expression. In stressed mice, vitamin B12 reduced DNAme in the Ttr promoter region. Importantly, using in vivo epigenome editing to alter DNAme in the brains of living mice for the first time, we establish a direct causal link between DNAme on Ttr and stress-associated behaviors. Discussion: In summary, using state-of-the-art techniques, this study uncovers a mechanistic link between cobalamin supplementation, Ttr, and markers of chronic stress and depression, encouraging further studies into dietary interventions for mood disorders.
2024-05-13 | GSE267047 | GEO
Project description:Background: Chronic stress significantly contributes to mood- and anxiety disorders. Previous data suggest a correlative connection between vitamin B12 supplementation, depression, and stress resilience. However, the underlying mechanisms are still poorly understood. Methods: Using the chronic variable stress mouse model coupled with RNA-sequencing, we determined vitamin B12-induced transcriptional changes related to stress resilience. By viral-mediated gene transfer and in vivo epigenome editing, we reveal a functional pathway linking vitamin B12, DNA methylation, and depressive-like symptoms. Results: We identified Transthyretin (Ttr) as a sex-specific key target of vitamin B12 action in chronic stress. Accordingly, TTR expression was increased postmortem in the prefrontal cortex of male, but not female, depressed patients. Virally altered Ttr in the prefrontal cortex functionally contributed to stress- and depression-related behaviors, changes in dendritic spine morphology, and gene expression. In stressed mice, vitamin B12 reduced DNAme in the Ttr promoter region. Importantly, using in vivo epigenome editing to alter DNAme in the brains of living mice for the first time, we establish a direct causal link between DNAme on Ttr and stress-associated behaviors. Discussion: In summary, using state-of-the-art techniques, this study uncovers a mechanistic link between cobalamin supplementation, Ttr, and markers of chronic stress and depression, encouraging further studies into dietary interventions for mood disorders.
2024-05-18 | GSE267339 | GEO
Project description:Ageing and mutations of transthyretin (TTR), the thyroid hormones and retinol transporting protein lead to amyloidosis by destabilizing the structure of TTR. Because protein structure is regulated through posttranslational modifications, we investigated the Small Ubiquitin-like Modifier (SUMO)ylation of TTR. We chose the widely used Ubc9 fusion-directed SUMOylation system, which is based on a fusion of the SUMOylation substrate of interest with Ubc9, a sole SUMO conjugating enzyme. Surprisingly, despite our presumptions, we found that Ubc9 fused to TTR was SUMOylated at a unique set of lysine residues. Three unknown SUMOylation sites of Ubc9—K154, K18 and K65—were revealed by mass spectrometry (MS). The previously reported SUMOylation at K49 of Ubc9 was also observed. SUMOylation of the lysine residues of TTR fused to Ubc9 was hardly detectable. However, non-fused TTR was SUMOylated via trans-SUMOylation by Ubc9 fused to TTR. Interestingly, mutating the catalytic residue of Ubc9 fused to TTR did not result in complete loss of the SUMOylation signal, suggesting that Ubc9 linked to TTR is directly cross-SUMOylated by the SUMO-activating enzyme E1. Ubc9, TTR or fusion proteins composed of TTR and Ubc9 specifically affected the global SUMOylation of cellular proteins. TTR or Ubc9 alone increased global SUMOylation, whereas TTR and Ubc9 together decreased the amount of high-molecular weight (HMW) SUMO conjugates. Our data suggest that TTR may influence the SUMOylation of Ubc9, thereby altering signalling pathways in the cell.
2016-08-15 | PXD004515 | Pride