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Molecular analysis, pathogenic mechanisms, and readthrough therapy on a large cohort of Kabuki syndrome patients.

ABSTRACT: Kabuki syndrome (KS) is a multiple congenital anomalies syndrome characterized by characteristic facial features and varying degrees of mental retardation, caused by mutations in KMT2D/MLL2 and KDM6A/UTX genes. In this study, we performed a mutational screening on 303 Kabuki patients by direct sequencing, MLPA, and quantitative PCR identifying 133 KMT2D, 62 never described before, and four KDM6A mutations, three of them are novel. We found that a number of KMT2D truncating mutations result in mRNA degradation through the nonsense-mediated mRNA decay, contributing to protein haploinsufficiency. Furthermore, we demonstrated that the reduction of KMT2D protein level in patients' lymphoblastoid and skin fibroblast cell lines carrying KMT2D-truncating mutations affects the expression levels of known KMT2D target genes. Finally, we hypothesized that the KS patients may benefit from a readthrough therapy to restore physiological levels of KMT2D and KDM6A proteins. To assess this, we performed a proof-of-principle study on 14 KMT2D and two KDM6A nonsense mutations using specific compounds that mediate translational readthrough and thereby stimulate the re-expression of full-length functional proteins. Our experimental data showed that both KMT2D and KDM6A nonsense mutations displayed high levels of readthrough in response to gentamicin treatment, paving the way to further studies aimed at eventually treating some Kabuki patients with readthrough inducers.

SUBMITTER: Micale L

PROVIDER: S-EPMC4234006 | biostudies-literature | 2014 Jul

REPOSITORIES: biostudies-literature

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Molecular analysis, pathogenic mechanisms, and readthrough therapy on a large cohort of Kabuki syndrome patients.

Micale Lucia L Augello Bartolomeo B Maffeo Claudia C Selicorni Angelo A Zucchetti Federica F Fusco Carmela C De Nittis Pasquelena P Pellico Maria Teresa MT Mandriani Barbara B Fischetto Rita R Boccone Loredana L Silengo Margherita M Biamino Elisa E Perria Chiara C Sotgiu Stefano S Serra Gigliola G Lapi Elisabetta E Neri Marcella M Ferlini Alessandra A Cavaliere Maria Luigia ML Chiurazzi Pietro P Monica Matteo Della MD Scarano Gioacchino G Faravelli Francesca F Ferrari Paola P Mazzanti Laura L Pilotta Alba A Patricelli Maria Grazia MG Bedeschi Maria Francesca MF Benedicenti Francesco F Prontera Paolo P Toschi Benedetta B Salviati Leonardo L Melis Daniela D Di Battista Eliana E Vancini Alessandra A Garavelli Livia L Zelante Leopoldo L Merla Giuseppe G

Human mutation 20140409 7

Kabuki syndrome (KS) is a multiple congenital anomalies syndrome characterized by characteristic facial features and varying degrees of mental retardation, caused by mutations in KMT2D/MLL2 and KDM6A/UTX genes. In this study, we performed a mutational screening on 303 Kabuki patients by direct sequencing, MLPA, and quantitative PCR identifying 133 KMT2D, 62 never described before, and four KDM6A mutations, three of them are novel. We found that a number of KMT2D truncating mutations result in mR ...[more]

PMID: 24633898

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Project description:BackgroundKabuki syndrome is a genetic disorder that affects several body systems and presents with variations in symptoms and severity. The syndrome is named for a common phenotype of faces resembling stage makeup used in a Japanese traditional theatrical art named kabuki. The most frequent cause of this syndrome is mutations in the H3K4 family of histone methyltransferases while a smaller percentage results from genetic alterations affecting the histone demethylase, KDM6A. Because of the rare presentation of the latter form of the disease, little is known about how missense changes in the KDM6A protein sequence impact protein function.ResultsIn this study, we use molecular mechanic and molecular dynamic simulations to enhance the annotation and mechanistic interpretation of the potential impact of eleven KDM6A missense variants found in Kabuki syndrome patients. These variants (N910S, D980V, S1025G, C1153R, C1153Y, P1195L, L1200F, Q1212R, Q1248R, R1255W, and R1351Q) are predicted to be pathogenic, likely pathogenic or of uncertain significance by sequence-based analysis. Here, we demonstrate, for the first time, that although Kabuki syndrome missense variants are found outside the functionally critical regions, they could affect overall function by significantly disrupting global and local conformation (C1153R, C1153Y, P1195L, L1200F, Q1212R, Q1248R, R1255W and R1351Q), chemical environment (C1153R, C1153Y, P1195L, L1200F, Q1212R, Q1248R, R1255W and R1351Q), and/or molecular dynamics of the catalytic domain (all variants). In addition, our approaches predict that many mutations, in particular C1153R, could allosterically disrupt the key enzymatic interactions of KDM6A.ConclusionsOur study demonstrates that the KDM6A Kabuki syndrome variants may impair histone demethylase function through various mechanisms that include altered protein integrity, local environment, molecular interactions and protein dynamics. Molecular dynamics simulations of the wild type and the variants are critical to gain a better understanding of molecular dysfunction. This type of comprehensive structure- and MD-based analyses should help develop improved impact scoring systems to interpret the damaging effects of variants in this protein and other related proteins as well as provide detailed mechanistic insight that is not currently predictable from sequence alone.

Project description:Kabuki syndrome (KS) is a well-recognized disorder characterized by postnatal growth deficiency, dysmorphic facial features, skeletal anomalies, and intellectual disability. The syndrome is caused by KMT2D gene mutations or less frequently KDM6A gene mutations or deletions. We report a systematic evaluation of KS patients from Campania region of Italy; data were also compared with literature ones. We collected data of 15 subjects (8 males and 7 females with age range 10-26 years; mean age 16.9 years) with confirmed diagnosis of KS, representing the entire cohort of patients from Campania Region. Each patient performed biochemical testing and instrumental investigation. Neuro-intellectual development, cranio-facial dysmorphisms, and multisystem involvement data were collected retrospectively. For each category, type of defects and frequency of the anomalies were analyzed. Our observation shows that KS patients from Campania region have some particular and previously underscored, neurological and immunological findings. We found high prevalence of EEG's abnormalities (43%) and MRI brain abnormalities (60%). Microcephaly resulted more common in our series (33%), if compared with major cohorts described in literature. Biochemical features of immunodeficiency and autoimmune diseases including thyroid autoimmunity, polyserositis, and vitiligo were observed with high prevalence (54.5%). Low immunoglobulins levels were a frequent finding. Lymphocyte class investigation showed significantly reduced CD8 levels in one patient.Conclusions: These data confirm great heterogeneity of clinical manifestations in KS and suggest to introduce further clinical diagnostic criteria in order to perform a correct and precocious diagnosis. What is Known • Kabuki syndrome is characterized by growth deficiency, dysmorphic facial features, skeletal anomalies, and intellectual disability • Immune dysfunction is a common finding but autoimmune diseases are rarely seen • Neurological features are common What is New • Some particular facial features could help gestalt diagnosis (hypertelorism, broad nasal bridge, micrognathia, tooth agenesis, cutaneous haemangiomas and strabismus) • Higher prevalence of autoimmune disorders than previously reported • Particular neurological features are present in this cohort (EEG and MRI brain abnormalities).

Project description:Vascular Ehlers-Danlos syndrome (vEDS) is a severe connective tissue disorder caused by dominant mutations in the COL3A1 gene, which encodes type III collagen (COLLIII). COLLIII is primarily found in blood vessels and hollow organs, and its deficiency causes fragile soft connective tissues, with life-threatening arterial and organ ruptures. There are currently no targeted therapies available. Although disease results from COLLIII misfolding caused by triple helix structure disruption, the underlying pathomechanisms are largely unknown. To address this knowledge gap, we conducted a comprehensive transcriptome analysis using RNA-seq on a large cohort of dermal fibroblasts from vEDS patients and healthy donors. Our investigation revealed an intricate interplay between proteostasis abnormalities, inefficient endoplasmic reticulum (ER) stress response, and compromised autophagy, which may significantly impact the molecular pathology. We reported the first comprehensive map of complex gene expression networks, which may provide significant molecular insights into intricate interactions within cellular signaling pathways. the current RNA-seq data not only confirm these initial findings but also provide convincing evidence that ineffective proteostasis, unbalanced ER homeostasis, and compromised autophagy could substantially impact the molecular pathology driving the disease. aken together, or findings suggest that the accumulation of these misfolded aggregates can impair critical cellular functions such as ER proteostasis, adaptative responses to counteract ER stress (UPR and ERAD pathways), autophagy, mTOR pathway, and lysosome function. Perturbation of these key biological processes can have an impact on a range of cell activities, including differentiation, proliferation, and apoptosis, all of which are associated with the pathophysiology of vEDS. This work was supported by Fondazione Telethon and the “Associazione con Giacomo contro vEDS”, (Grant number: GSA21F001, Seed Grant Fall 2021 vEDS) to Nicola Chiarelli.

Dataset Information

Molecular analysis, pathogenic mechanisms, and readthrough therapy on a large cohort of Kabuki syndrome patients.

Publications

Molecular analysis, pathogenic mechanisms, and readthrough therapy on a large cohort of Kabuki syndrome patients.

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