Bulk RNA-sequencing from whole heart lysates of a mouse model of hypertrophic cardiomyopathy after TTL overexpression.
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE233851 | GEO | 2024/08/27
REPOSITORIES: GEO
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Project description:Rationale Hypertrophic cardiomyopathy (HCM) is the most common cardiac genetic disorder caused by sarcomeric gene variants and is associated with left ventricular hypertrophy and diastolic dysfunction. The role of the microtubule network has recently gained interest with findings that microtubule detyrosination (dTyr-MTs) is markedly elevated in heart failure. Acute reduction of dTyr-MTs by inhibition of the detyrosinase (VASH/SVBP complex) or activation of the tyrosinase (tubulin tyrosine ligase, TTL) markedly improved contractility and reduced stiffness in human failing cardiomyocytes, presenting a new perspective for HCM treatment. Objective In this study, we tested the impact of chronic tubulin tyrosination in a HCM mouse model (Mybpc3-knock-in; KI), in human HCM cardiomyocytes, and in SVBP-deficient human engineered heart tissues (EHTs). Methods and Results AAV9-mediated TTL transfer was applied in neonatal wild-type (WT) rodents, in 3-week-old KI mice, and in HCM human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes. We show: i) TTL for 6 weeks dose-dependently reduced dTyr-MTs and improved contractility without affecting cytosolic calcium transients in WT cardiomyocytes. ii) TTL for 12 weeks reduced the abundance of dTyr-MTs in the myocardium, improved diastolic filling, compliance, cardiac output, and stroke volume in KI mice. iii) TTL for 10 days normalized cell area in HCM hiPSC-cardiomyocytes. iv) TTL overexpression activates transcription of several tubulin isoforms, and omics GO analysis revealed enrichment of components of the cytoskeleton, sarcomere Z-disc, mitochondria, and intercalated disc in KI mice. v) SVBP-deficient EHTs exhibited reduced dTyr-MT levels, higher force, and faster relaxation than TTL-deficient and WT EHTs. RNA-seq and mass spectrometry analysis revealed distinct enrichment of cardiomyocyte components and pathways in SVBP-KO vs. TTL-KO EHTs. Conclusion This study provides the first proof-of-concept that chronic activation of tubulin tyrosination in HCM mice and in human EHTs improves heart function and holds promise for targeting the non-sarcomeric cytoskeleton in heart disease.
2024-08-28 | PXD054236 | Pride
Project description:BACKGROUND: MYBPC3 is one of the most mutated gene known to cause hypertrophic cardiomyopathy (HCM). However, the molecular mechanisms of how mutations in MYBPC3 lead to the onset and progression of HCM are poorly understood. Thus, advancing in-vitro studies to define these mechanisms of mutations leading to HCM are still warranted. Thus, the primary objective of this study was to investigate the molecular mechanisms underlying the pathogenesis of HCM associated with MYBPC3 mutation utilizing isogenic human-induced pluripotent stem cell (hiPSC)-derived cardiac organoids (hCOs).
2025-01-28 | GSE262876 | GEO
Project description:Tubulin detyrosination-tyrosination cycle regulates the stability of microtubules. Thus far described on α-tubulins, the tyrosination level is maintained by a single tubulin-tyrosine ligase (TTL). However, the precise dynamics and tubulin isoforms which undergo (de)tyrosination in neurons are unknown. Here, we exploit the substrate promiscuity of the TTL to introduce an O-propargyl-L-tyrosine in cancer cell lines and neurons. Mass spectrometry-based chemical proteomics in neuroblastoma cells using the O-propargyl-L-tyrosine probe revealed previously discussed tyrosination of TUBA4A, MAPRE1, and other non-tubulin proteins. This finding was further corroborated in differentiating neurons. We present the method for tubulin tyrosination profiling in living cells. Our results show that detyrosination-tyrosination is not restricted to α-tubulins with coded C-terminal tyrosine and is thus involved in fine-tuning of the tubulin and non-tubulin proteins during neuronal differentiation.
2022-10-20 | PXD037402 | Pride
Project description:Hypertrophic cardiomyopathy (HCM) is an inherited cardiac disorder characterized by ventricular wall hypertrophy and diastolic dysfunction. Most patients with HCM are often a slowly progressive disease and may be asymptomatic; however, some progress to the dilated phase of HCM (D-HCM), and have a poorer prognosis. This study established disease-specific human induced pluripotent stem cells (iPSCs) from a patient with D-HCM harboring a truncating mutation in MYBPC3, and compared transcriptomes of iPSCs-derived cardiomyocytes between healthy control and D-HCM.
2025-04-06 | GSE263980 | GEO
Project description:Post-translational cycles of α-tubulin detyrosination/tyrosination generate microtubule diversity, with broad physiological and clinical implications. However, the mechanistic understanding of how this microtubule diversity is decoded into specific cellular functions remains largely unknown. Here we show that the core microtubule-binding site at kinetochores ‘reads’ and responds to the α-tubulin detyrosination/tyrosination code. By combining CRISPR-Cas9 gene editing of enzymatic ‘writers’ of α-tubulin detyrosination/tyrosination in human cells, with biochemical purifications, mass spectrometry and super-resolution microscopy, we identified over a hundred proteins that preferentially associate with either detyrosinated or tyrosinated microtubules during mitosis. Among these, the conserved KNL-1/MIS12 complex/NDC80 complex (KMN) network at kinetochores showed a preference for tyrosinated microtubules. In addition to established roles in error correction and polar chromosome alignment, live-cell imaging uncovered a previously overlooked link between the α-tubulin detyrosination/tyrosination code and the formation of functional kinetochore-microtubule attachments, necessary for normal metaphase chromosome oscillations and timely spindle assembly checkpoint satisfaction. In vitro reconstitution experiments revealed that α-tubulin detyrosination specifically promotes Ndc80 diffusion along the microtubule lattice. We propose that the gradual accumulation of detyrosinated α-tubulin after initial microtubule capture at kinetochores facilitates the biased diffusion of core-binding proteins along the microtubule lattice, establishing a positive feedback loop that sustains kinetochore-microtubule attachments.
2024-10-03 | PXD047133 | Pride
Project description:Myosin inhibitor mavacamten is the first targeted treatment available for hypertrophic cardiomyopathy (HCM), a disease caused by hundreds of genetic variants that affect mainly sarcomeric myosin and its negative regulator cardiac myosin-binding protein C (cMyBP-C, encoded by MYBPC3). Here, we have examined whether the reported limited efficacy of mavacamten in a fraction of HCM patients can result from dissimilar HCM pathomechanisms triggered by different genetic variants, a scenario particularly relevant for MYBPC3-associated HCM. To this aim, we have generated knock-in mice including missense pathogenic variant cMyBP-C p.R502W, which, different from patients who carry truncations in the protein, develop progressive pathogenic myocardial remodeling in the absence of alterations of cMyBP-C levels and localization. Mechanistically, mutation R502W reduces the binding affinity of cMyBP-C for myosin, increases Ca2+ sensitivity and promotes the ON structural state of myosin heads without inducing a shift towards more active myosin conformations as observed when cMyBP-C levels are reduced. Despite these differences, we demonstrate that mavacamten blunts myocardial remodeling both in R502W and cMyBP-C-deficient, knock-out hearts, correlating with its ability to normalize the levels of ON myosins in R502W sarcomeres. These beneficial effects are accompanied by improved tolerance to exercise in R502W animals. In human engineered heart tissues carrying R502W, we show that mavacamten opposes hypercontractility induced by the mutation. Hence, in combination our results indicate that myosin inhibition is effective to treat HCM caused by both truncating and missense variants in MYBPC3 regardless of the primary pathomechanisms they elicit.
2026-06-02 | GSE326720 | GEO
Project description:Myosin inhibitor mavacamten is the first targeted treatment available for hypertrophic cardiomyopathy (HCM), a disease caused by hundreds of genetic variants that affect mainly sarcomeric myosin and its negative regulator cardiac myosin-binding protein C (cMyBP-C, encoded by MYBPC3). Here, we have examined whether the reported limited efficacy of mavacamten in a fraction of HCM patients can result from dissimilar HCM pathomechanisms triggered by different genetic variants, a scenario particularly relevant for MYBPC3-associated HCM. To this aim, we have generated knock-in mice including missense pathogenic variant cMyBP-C p.R502W, which, different from patients who carry truncations in the protein, develop progressive pathogenic myocardial remodeling in the absence of alterations of cMyBP-C levels and localization. Mechanistically, mutation R502W reduces the binding affinity of cMyBP-C for myosin, increases Ca2+ sensitivity and promotes the ON structural state of myosin heads without inducing a shift towards more active myosin conformations as observed when cMyBP-C levels are reduced. Despite these differences, we demonstrate that mavacamten blunts myocardial remodeling both in R502W and cMyBP-C-deficient, knock-out hearts, correlating with its ability to normalize the levels of ON myosins in R502W sarcomeres. These beneficial effects are accompanied by improved tolerance to exercise in R502W animals. In human engineered heart tissues carrying R502W, we show that mavacamten opposes hypercontractility induced by the mutation. Hence, in combination our results indicate that myosin inhibition is effective to treat HCM caused by both truncating and missense variants in MYBPC3 regardless of the primary pathomechanisms they elicit.
2026-06-02 | GSE326721 | GEO
Project description:We report the application of RNA-sequencing technology for high-throughput profiling of Drosophila that express clinical variants of feline-MyBPC3 associated with Hypertrophic Cardiomyopathy (HCM).
2020-12-04 | GSE141574 | GEO
Project description:Myosin inhibitor mavacamten is the first targeted treatment available for hypertrophic cardiomyopathy (HCM), a disease caused by hundreds of genetic variants that affect mainly sarcomeric myosin and its negative regulator cardiac myosin-binding protein C (cMyBP-C, encoded by MYBPC3). Here, we have examined whether the reported limited efficacy of mavacamten in a fraction of HCM patients can result from dissimilar HCM pathomechanisms triggered by different genetic variants, a scenario particularly relevant for MYBPC3-associated HCM. To this aim, we have generated knock-in mice including missense pathogenic variant cMyBP-C p.R502W, which, different from patients who carry truncations in the protein, develop progressive pathogenic myocardial remodeling in the absence of alterations of cMyBP-C levels and localization. Mechanistically, mutation R502W reduces the binding affinity of cMyBP-C for myosin, increases Ca2+ sensitivity and promotes the ON structural state of myosin heads without inducing a shift towards more active myosin conformations as observed when cMyBP-C levels are reduced. Despite these differences, we demonstrate that mavacamten blunts myocardial remodeling both in R502W and cMyBP-C-deficient, knock-out hearts, correlating with its ability to normalize the levels of ON myosins in R502W sarcomeres. These beneficial effects are accompanied by improved tolerance to exercise in R502W animals. In human engineered heart tissues carrying R502W, we show that mavacamten opposes hypercontractility induced by the mutation. Hence, in combination our results indicate that myosin inhibition is effective to treat HCM caused by both truncating and missense variants in MYBPC3 regardless of the primary pathomechanisms they elicit.
2026-06-02 | PXD076207 | Pride
Project description:The study investigates the effect of a heterozygous MYBPC3 mutation combined with Western diet feeding on protein expression in a HCM mouse model. Hypertrophic cardiomyopathy (HCM) is the most common inherited genetic heart disease, characterized by asymmetric left ventricular hypertrophy and diastolic dysfunction. It is generally accepted that HCM is caused by mutations in genes encoding sarcomere proteins (e.g. MYBPC3, MYH7). However, not all mutation carriers will develop HCM and, moreover, phenotypical variation in terms of clinical presentation is large. The phenotypical expression of HCM thus requires additional disease modifiers on top of a sarcomere gene mutation. Clinical reports identify obesity and related comorbidities (diabetes, hyperlipidemia, hypertension) as a major factor contributing to disease expression and severity. We aimed to mimic this situation in a HCM mouse model by feeding a Western diet (8 weeks) to mice carrying a heterozygous mutation in MYBPC3. These heterozygous mice are phenotype negative under normal conditions, but developed cardiac dysfunction and hypertrophy upon Western diet feeding. The main goal of this proteomic screening is to identify clusters of proteins and pathways that are down/upregulated specifically in the left ventricle of mice suffering from a mutation and Western diet feeding. Also, we are interested in protein expression differences caused solely by mutation or Western diet. Group 1 are Wild type mice that received normal chow; group 2 are heterozygous mice that received normal chow; group 3 are Wild type mice that received a Western diet; group 4 are heterozygous mice that received a Western diet.
2023-10-01 | PXD043345 | Pride