Molecular Analysis, Pathogenic Mechanisms, and Readthrough Therapy on a Large Cohort of Kabuki Syndrome Patients

Lucia Micale, Bartolomeo Augello, Claudia Maffeo, Angelo Selicorni, Federica Zucchetti, Carmela Fusco, Pasquelena De Nittis, Maria Teresa Pellico, Barbara Mandriani, Rita Fischetto, Loredana Boccone, Margherita Silengo, Elisa Biamino, Chiara Perria, Stefano Sotgiu, Gigliola Serra, Elisabetta Lapi, Marcella Neri, Alessandra Ferlini, Maria Luigia CavalierePietro Chiurazzi, Matteo Della Monica, Gioacchino Scarano, Francesca Faravelli, Paola Ferrari, Laura Mazzanti, Alba Pilotta, Maria Grazia Patricelli, Maria Francesca Bedeschi, Francesco Benedicenti, Paolo Prontera, Benedetta Toschi, Leonardo Salviati, Daniela Melis, Eliana Di Battista, Alessandra Vancini, Livia Garavelli, Leopoldo Zelante, Giuseppe Merla

Research output: Contribution to journalArticle

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. In this report, we have expanded the spectrum of mutations of KMT2D and KDM6A genes by analysing our cohort of 303 Kabuki patients by direct sequencing, MLPA and quantitative PCR. Based on KMT2D biological role, we designed functional studies that highlighted the haploinsufficiency of KMT2D as one of the mechanisms underlying the pathogenesis of the disease. Moreover, we provided the first preliminary proof-of-concept that occurring nonsense mutations in KMT2D and KDM6A can be effectively suppressed and the functional endogenous protein level and biological activity of KMT2D and KDM6A proteins restored.

Original languageEnglish
Pages (from-to)841-850
Number of pages10
JournalHuman Mutation
Volume35
Issue number7
DOIs
Publication statusPublished - 2014

Keywords

  • Haploinsufficiency
  • Kabuki syndrome
  • KDM6A
  • KMT2D
  • Readthrough

ASJC Scopus subject areas

  • Genetics
  • Genetics(clinical)

Fingerprint Dive into the research topics of 'Molecular Analysis, Pathogenic Mechanisms, and Readthrough Therapy on a Large Cohort of Kabuki Syndrome Patients'. Together they form a unique fingerprint.

  • Cite this

    Micale, L., Augello, B., Maffeo, C., Selicorni, A., Zucchetti, F., Fusco, C., De Nittis, P., Pellico, M. T., Mandriani, B., Fischetto, R., Boccone, L., Silengo, M., Biamino, E., Perria, C., Sotgiu, S., Serra, G., Lapi, E., Neri, M., Ferlini, A., ... Merla, G. (2014). Molecular Analysis, Pathogenic Mechanisms, and Readthrough Therapy on a Large Cohort of Kabuki Syndrome Patients. Human Mutation, 35(7), 841-850. https://doi.org/10.1002/humu.22547