ALPK3-deficient cardiomyocytes generated from patient-derived induced pluripotent stem cells and mutant human embryonic stem cells display abnormal calcium handling and establish that ALPK3 deficiency underlies familial cardiomyopathy.
We identified a novel homozygous truncating mutation in the gene encoding alpha kinase 3 (ALPK3) in a family presenting with paediatric cardiomyopathy. A recent study identified biallelic truncating mutations of ALPK3 in three unrelated families; therefore, there is strong genetic evidence that ALPK3 mutation causes cardiomyopathy. This study aimed to clarify the mutation mechanism and investigate the molecular and cellular pathogenesis underlying ALPK3-mediated cardiomyopathy.



We performed detailed clinical and genetic analyses of a consanguineous family, identifying a new ALPK3 mutation (c.3792G>A, p.W1264X) which undergoes nonsense-mediated decay in ex vivo and in vivo tissues. Ultra-structural analysis of cardiomyocytes derived from patient-specific and human ESC-derived stem cell lines lacking ALPK3 revealed disordered sarcomeres and intercalated discs. Multi-electrode array analysis and calcium imaging demonstrated an extended field potential duration and abnormal calcium handling in mutant contractile cultures.



This study validates the genetic evidence, suggesting that mutations in ALPK3 can cause familial cardiomyopathy and demonstrates loss of function as the underlying genetic mechanism. We show that ALPK3-deficient cardiomyocytes derived from pluripotent stem cell models recapitulate the ultrastructural and electrophysiological defects observed in vivo. Analysis of differentiated contractile cultures identified abnormal calcium handling as a potential feature of cardiomyocytes lacking ALPK3, providing functional insights into the molecular mechanisms underlying ALPK3-mediated cardiomyopathy.
Bruce Lefroy Centre for Genetic Health Research, Murdoch Childrens Research Institute, Flemington Road, Parkville 3052, Victoria, Australia Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, The University of Melbourne, Parkville 3052, Victoria, Australia.
Dean G
DG
Phelan
Murdoch Childrens Research Institute, Royal Children's Hospital, Flemington Road, Parkville 3052, Victoria, Australia.
David J
DJ
Anderson
Murdoch Childrens Research Institute, Royal Children's Hospital, Flemington Road, Parkville 3052, Victoria, Australia Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, The University of Melbourne, Parkville 3052, Victoria, Australia.
Sara E
SE
Howden
Centre for Eye Research Australia, Royal Victorian Eye and Ear Hospital; Ophthalmology, Department of Surgery, University of Melbourne, 32 Gisborne Street, East Melbourne 3002, Victoria, Australia.
Raymond C B
RC
Wong
Population Health and Immunity Division, Walter and Eliza Hall Institute, 1G Royal Parade, Melbourne 3052, Victoria, Australia.
Peter F
PF
Hickey
Bruce Lefroy Centre for Genetic Health Research, Murdoch Childrens Research Institute, Flemington Road, Parkville 3052, Victoria, Australia.
Kate
K
Pope
Bruce Lefroy Centre for Genetic Health Research, Murdoch Childrens Research Institute, Flemington Road, Parkville 3052, Victoria, Australia Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, The University of Melbourne, Parkville 3052, Victoria, Australia.
Gabrielle R
GR
Wilson
Centre for Eye Research Australia, Royal Victorian Eye and Ear Hospital; Ophthalmology, Department of Surgery, University of Melbourne, 32 Gisborne Street, East Melbourne 3002, Victoria, Australia.
Alice
A
Pébay
Murdoch Childrens Research Institute, Royal Children's Hospital, Flemington Road, Parkville 3052, Victoria, Australia Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, The University of Melbourne, Parkville 3052, Victoria, Australia Department of Cardiology, The Royal Children's Hospital, Parkville 3052, Victoria, Australia.
Andrew M
AM
Davis
The Florey Institute for Neuroscience and Mental Health, The University of Melbourne, Parkville 3052, Victoria, Australia.
Steven
S
Petrou
Murdoch Childrens Research Institute, Royal Children's Hospital, Flemington Road, Parkville 3052, Victoria, Australia Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, The University of Melbourne, Parkville 3052, Victoria, Australia Department of Anatomy and Developmental Biology, Faculty of Medicine, Nursing and Health Sciences, Monash University, Clayton 3800, Victoria, Australia.
Andrew G
AG
Elefanty
Murdoch Childrens Research Institute, Royal Children's Hospital, Flemington Road, Parkville 3052, Victoria, Australia Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, The University of Melbourne, Parkville 3052, Victoria, Australia Department of Anatomy and Developmental Biology, Faculty of Medicine, Nursing and Health Sciences, Monash University, Clayton 3800, Victoria, Australia.
Edouard G
EG
Stanley
Genetic Medicine, Royal Melbourne Hospital, Parkville 3052, Victoria, Australia.
Paul A
PA
James
Victorian Clinical Genetics Services, Murdoch Childrens Research Institute, Flemington Road, Parkville 3052, Victoria, Australia.
Ivan
I
Macciocca
Population Health and Immunity Division, Walter and Eliza Hall Institute, 1G Royal Parade, Melbourne 3052, Victoria, Australia Department of Medical Biology, The University of Melbourne, Parkville 3052, Victoria, Australia.
Melanie
M
Bahlo
Department of Cardiology, The Royal Children's Hospital, Parkville 3052, Victoria, Australia Murdoch Childrens Research Institute, Royal Children's Hospital, Flemington Road, Parkville 3052, Victoria, Australia Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, The University of Melbourne, Parkville 3052, Victoria, Australia.
Michael M
MM
Cheung
Bruce Lefroy Centre for Genetic Health Research, Murdoch Childrens Research Institute, Flemington Road, Parkville 3052, Victoria, Australia Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, The University of Melbourne, Parkville 3052, Victoria, Australia Victorian Clinical Genetics Services, Murdoch Childrens Research Institute, Flemington Road, Parkville 3052, Victoria, Australia.
David J
DJ
Amor
Murdoch Childrens Research Institute, Royal Children's Hospital, Flemington Road, Parkville 3052, Victoria, Australia School of Biosciences, The University of Melbourne, Parkville 3052, Victoria, Australia david.elliott@mcri.edu.au paul.lockhart@mcri.edu.au.
David A
DA
Elliott
Bruce Lefroy Centre for Genetic Health Research, Murdoch Childrens Research Institute, Flemington Road, Parkville 3052, Victoria, Australia Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, The University of Melbourne, Parkville 3052, Victoria, Australia david.elliott@mcri.edu.au paul.lockhart@mcri.edu.au.
Paul J
PJ
Lockhart
D002118:Calcium; D009202:Cardiomyopathies; D000066449:Human Embryonic Stem Cells; D006801:Humans; D057026:Induced Pluripotent Stem Cells; D009124:Muscle Proteins; D032383:Myocytes, Cardiac; D011494:Protein Kinases
D002118:Calcium; D009124:Muscle Proteins; D011494:Protein Kinases; C000618184:Alpk3 protein, human
Calcium handling; Intercalated disc; Stem cells; alpha kinase 3