NKX2-5 mutations causative for congenital heart disease retain functionality and are directed to hundreds of targets
Romaric Bouveret,
Ashley J Waardenberg,
Nicole Schonrock,
Mirana Ramialison,
Tram Doan,
Danielle de Jong,
Antoine Bondue,
Gurpreet Kaur,
Stephanie Mohamed,
Hananeh Fonoudi,
Chiann-mun Chen,
Merridee A Wouters,
Shoumo Bhattacharya,
Nicolas Plachta,
Sally L Dunwoodie,
Gavin Chapman,
Cédric Blanpain,
Richard P Harvey
Affiliations
Romaric Bouveret
Victor Chang Cardiac Research Institute, Darlinghurst, Australia; St. Vincent's Clinical School, University of New South Wales, Sydney, Australia
Ashley J Waardenberg
Victor Chang Cardiac Research Institute, Darlinghurst, Australia; Children's Medical Research Institute, Sydney, Australia
Nicole Schonrock
Victor Chang Cardiac Research Institute, Darlinghurst, Australia; St. Vincent's Clinical School, University of New South Wales, Sydney, Australia
Mirana Ramialison
Victor Chang Cardiac Research Institute, Darlinghurst, Australia; St. Vincent's Clinical School, University of New South Wales, Sydney, Australia; European Molecular Biology Laboratory, Australian Regenerative Medicine Institute, Monash University, Clayton, Australia
Tram Doan
Victor Chang Cardiac Research Institute, Darlinghurst, Australia
Danielle de Jong
Victor Chang Cardiac Research Institute, Darlinghurst, Australia
Antoine Bondue
Institut de Recherche Interdisciplinaire en Biologie Humaine et Moléculaire, Université Libre de Bruxelles, Brussels, Belgium; Department of Cardiology, Erasme Hospital, Brussels, Belgium
Gurpreet Kaur
European Molecular Biology Laboratory, Australian Regenerative Medicine Institute, Monash University, Clayton, Australia
Stephanie Mohamed
Victor Chang Cardiac Research Institute, Darlinghurst, Australia
Hananeh Fonoudi
Victor Chang Cardiac Research Institute, Darlinghurst, Australia; St. Vincent's Clinical School, University of New South Wales, Sydney, Australia; Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, Academic Center for Education, Culture and Research, Tehran, Iran
Chiann-mun Chen
Department of Cardiovascular Medicine, Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, United Kingdom
Bioinformatics, Olivia Newton-John Cancer Research Institute, Melbourne, Australia; School of Life and Environmental Sciences, Deakin University, Geelong, Australia
Shoumo Bhattacharya
Department of Cardiovascular Medicine, Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, United Kingdom
Nicolas Plachta
European Molecular Biology Laboratory, Australian Regenerative Medicine Institute, Monash University, Clayton, Australia
Sally L Dunwoodie
Victor Chang Cardiac Research Institute, Darlinghurst, Australia; St. Vincent's Clinical School, University of New South Wales, Sydney, Australia; School of Biotechnology and Biomolecular Sciences, University of New South Wales, Kensington, Australia
Gavin Chapman
Victor Chang Cardiac Research Institute, Darlinghurst, Australia; St. Vincent's Clinical School, University of New South Wales, Sydney, Australia
Cédric Blanpain
Institut de Recherche Interdisciplinaire en Biologie Humaine et Moléculaire, Université Libre de Bruxelles, Brussels, Belgium; Walloon Excellence in Life Sciences and Biotechnology, Université Libre de Bruxelles, Brussels, Belgium
Richard P Harvey
Victor Chang Cardiac Research Institute, Darlinghurst, Australia; St. Vincent's Clinical School, University of New South Wales, Sydney, Australia
We take a functional genomics approach to congenital heart disease mechanism. We used DamID to establish a robust set of target genes for NKX2-5 wild type and disease associated NKX2-5 mutations to model loss-of-function in gene regulatory networks. NKX2-5 mutants, including those with a crippled homeodomain, bound hundreds of targets including NKX2-5 wild type targets and a unique set of "off-targets", and retained partial functionality. NKXΔHD, which lacks the homeodomain completely, could heterodimerize with NKX2-5 wild type and its cofactors, including E26 transformation-specific (ETS) family members, through a tyrosine-rich homophilic interaction domain (YRD). Off-targets of NKX2-5 mutants, but not those of an NKX2-5 YRD mutant, showed overrepresentation of ETS binding sites and were occupied by ETS proteins, as determined by DamID. Analysis of kernel transcription factor and ETS targets show that ETS proteins are highly embedded within the cardiac gene regulatory network. Our study reveals binding and activities of NKX2-5 mutations on WT target and off-targets, guided by interactions with their normal cardiac and general cofactors, and suggest a novel type of gain-of-function in congenital heart disease.
