Riboflavin-Responsive and -Non-responsive Mutations in FAD Synthase Cause Multiple Acyl-CoA Dehydrogenase and Combined Respiratory-Chain Deficiency.
Multiple acyl-CoA dehydrogenase deficiencies (MADDs) are a heterogeneous group of metabolic disorders with combined respiratory-chain deficiency and a neuromuscular phenotype. Despite recent advances in understanding the genetic basis of MADD, a number of cases remain unexplained. Here, we report clinically relevant variants in FLAD1, which encodes FAD synthase (FADS), as the cause of MADD and respiratory-chain dysfunction in nine individuals recruited from metabolic centers in six countries. In most individuals, we identified biallelic frameshift variants in the molybdopterin binding (MPTb) domain, located upstream of the FADS domain. Inasmuch as FADS is essential for cellular supply of FAD cofactors, the finding of biallelic frameshift variants was unexpected. Using RNA sequencing analysis combined with protein mass spectrometry, we discovered FLAD1 isoforms, which only encode the FADS domain. The existence of these isoforms might explain why affected individuals with biallelic FLAD1 frameshift variants still harbor substantial FADS activity. Another group of individuals with a milder phenotype responsive to riboflavin were shown to have single amino acid changes in the FADS domain. When produced in E. coli, these mutant FADS proteins resulted in impaired but detectable FADS activity; for one of the variant proteins, the addition of FAD significantly improved protein stability, arguing for a chaperone-like action similar to what has been reported in other riboflavin-responsive inborn errors of metabolism. In conclusion, our studies identify FLAD1 variants as a cause of potentially treatable inborn errors of metabolism manifesting with MADD and shed light on the mechanisms by which FADS ensures cellular FAD homeostasis.
Research Unit for Molecular Medicine, Department of Clinical Medicine, Aarhus University and University Hospital, 8200 Aarhus N, Denmark. Electronic address: rikke.olsen@clin.au.dk.
Rikke K J
RKJ
Olsen
Institute of Human Genetics, Technische Universität München, 81675 Munich, Germany; Institute of Human Genetics, Helmholtz Zentrum München, 85764 Neuherberg, Germany.
Eliška
E
Koňaříková
Department of Biosciences, Biotechnology, and Biopharmaceutics, University of Bari Aldo Moro, 70125 Bari, Italy.
Teresa A
TA
Giancaspero
Research Unit for Molecular Medicine, Department of Clinical Medicine, Aarhus University and University Hospital, 8200 Aarhus N, Denmark.
Signe
S
Mosegaard
Wellcome Trust Centre for Mitochondrial Research, Institute of Genetic Medicine, Newcastle University, Newcastle upon Tyne NE1 3BZ, UK.
Veronika
V
Boczonadi
Department of Paediatrics, Paracelsus Medical University, SALK Salzburg, 5020 Salzburg, Austria.
Lavinija
L
Mataković
Service Maladies Héréditaires du Métabolisme et Dépistage Néonatal, Centre de Biologie et Pathologie Est, Centre Hospitalier Universitaire Lyon, 69500 Bron, France.
Alice
A
Veauville-Merllié
Institute of Human Genetics, Helmholtz Zentrum München, 85764 Neuherberg, Germany.
Caterina
C
Terrile
Institute of Human Genetics, Technische Universität München, 81675 Munich, Germany; Institute of Human Genetics, Helmholtz Zentrum München, 85764 Neuherberg, Germany.
Thomas
T
Schwarzmayr
Institute of Human Genetics, Technische Universität München, 81675 Munich, Germany; Institute of Human Genetics, Helmholtz Zentrum München, 85764 Neuherberg, Germany.
Tobias B
TB
Haack
Clinical Neurosciences, Neurology, University of Helsinki and Helsinki University Hospital, 340 Helsinki, Finland.
Mari
M
Auranen
Department of Biosciences, Biotechnology, and Biopharmaceutics, University of Bari Aldo Moro, 70125 Bari, Italy.
Piero
P
Leone
Department DiBEST (Biology, Ecology, and Earth Sciences), University of Calabria, 87036 Arcavacata di Rende, Italy.
Michele
M
Galluccio
Biochemistry Hormonology Laboratory, Robert-Debré Hospital, 75019 Paris, France; Pharmacy Faculty, Paris Sud University, 92019 Chatenay-Malabry, France.
Apolline
A
Imbard
Wellcome Trust Centre for Mitochondrial Research, Institute of Genetic Medicine, Newcastle University, Newcastle upon Tyne NE1 3BZ, UK; Centro Andaluz de Biología del Desarrollo, Universidad Pablo de Olavide, 41013 Seville, Spain.
Purificacion
P
Gutierrez-Rios
Research Unit for Molecular Medicine, Department of Clinical Medicine, Aarhus University and University Hospital, 8200 Aarhus N, Denmark.
Johan
J
Palmfeldt
Institute of Human Genetics, Helmholtz Zentrum München, 85764 Neuherberg, Germany.
Elisabeth
E
Graf
Service Maladies Héréditaires du Métabolisme et Dépistage Néonatal, Centre de Biologie et Pathologie Est, Centre Hospitalier Universitaire Lyon, 69500 Bron, France.
Christine
C
Vianey-Saban
Neurometabolic Unit, National Hospital for Neurology and Neurosurgery, London WCIN 3BG, UK.
Marcus
M
Oppenheim
INSERM UMR 1141, Hôpital Robert Debré, 75019 Paris, France; Reference Center for Inherited Metabolic Diseases, Robert-Debré Hospital, Assistance Publique - Hôpitaux de Paris, 75019 Paris, France; Faculté de Médecine Denis Diderot, Université Paris Diderot (Paris 7), 75013 Paris, France.
Manuel
M
Schiff
Reference Center for Inherited Metabolic Diseases, Robert-Debré Hospital, Assistance Publique - Hôpitaux de Paris, 75019 Paris, France.
Samia
S
Pichard
Biochemistry Hormonology Laboratory, Robert-Debré Hospital, 75019 Paris, France.
Odile
O
Rigal
Wellcome Trust Centre for Mitochondrial Research, Institute of Genetic Medicine, Newcastle University, Newcastle upon Tyne NE1 3BZ, UK.
Angela
A
Pyle
Wellcome Trust Centre for Mitochondrial Research, Institute of Genetic Medicine, Newcastle University, Newcastle upon Tyne NE1 3BZ, UK; Department of Clinical Neurosciences, Cambridge Biomedical Campus, University of Cambridge, Cambridge CB2 0QQ, UK.
Patrick F
PF
Chinnery
Department of Pediatrics, Medical University of Vienna, 1090 Vienna, Austria.
Vassiliki
V
Konstantopoulou
Department of Pediatrics, Medical University of Vienna, 1090 Vienna, Austria.
Dorothea
D
Möslinger
Department of Paediatrics, Paracelsus Medical University, SALK Salzburg, 5020 Salzburg, Austria.
René G
RG
Feichtinger
Pathology Unit, Department of Pediatrics, Hacettepe University Children's Hospital, 06100 Ankara, Turkey.
Beril
B
Talim
Neurology Unit, Department of Pediatrics, Hacettepe University Children's Hospital, 06100 Ankara, Turkey.
Haluk
H
Topaloglu
Metabolism Unit, Department of Pediatrics, Hacettepe University Children's Hospital, 06100 Ankara, Turkey.
Turgay
T
Coskun
Pathology Unit, Department of Pediatrics, Hacettepe University Children's Hospital, 06100 Ankara, Turkey.
Safak
S
Gucer
Medical Genetics Section, Department of Biomedicine and Prevention, Tor Vergata University of Rome, 00133 Rome, Italy.
Annalisa
A
Botta
Neuromuscular Center, Department of Neurosciences, University of Padova, 35129 Padova, Italy.
Elena
E
Pegoraro
Neuromuscular Center, Department of Neurosciences, University of Padova, 35129 Padova, Italy.
Adriana
A
Malena
Neuromuscular Center, Department of Neurosciences, University of Padova, 35129 Padova, Italy.
Lodovica
L
Vergani
Italy Institute of Medical Genetics, Catholic University of Roma, 00168 Rome, Italy.
Daniela
D
Mazzà
Italy Institute of Medical Genetics, Catholic University of Roma, 00168 Rome, Italy.
Marcella
M
Zollino
Molecular Neurogenetics Unit, Foundation IRCCS Neurological Institute C. Besta, 20126 Milan, Italy.
Daniele
D
Ghezzi
Service Maladies Héréditaires du Métabolisme et Dépistage Néonatal, Centre de Biologie et Pathologie Est, Centre Hospitalier Universitaire Lyon, 69500 Bron, France.
Cecile
C
Acquaviva
Department of Pediatric Neurology, Hospital for Children and Adolescence, Helsinki University Central Hospital, 280 Helsinki, Finland.
Tiina
T
Tyni
Murdoch Childrens Research Institute and University of Melbourne, Melbourne, VIC 3010, Australia.
Avihu
A
Boneh
Institute of Human Genetics, Technische Universität München, 81675 Munich, Germany; Institute of Human Genetics, Helmholtz Zentrum München, 85764 Neuherberg, Germany.
Thomas
T
Meitinger
Institute of Human Genetics, Technische Universität München, 81675 Munich, Germany; Institute of Human Genetics, Helmholtz Zentrum München, 85764 Neuherberg, Germany.
Tim M
TM
Strom
Research Unit for Molecular Medicine, Department of Clinical Medicine, Aarhus University and University Hospital, 8200 Aarhus N, Denmark.
Niels
N
Gregersen
Department of Paediatrics, Paracelsus Medical University, SALK Salzburg, 5020 Salzburg, Austria.
Johannes A
JA
Mayr
Wellcome Trust Centre for Mitochondrial Research, Institute of Genetic Medicine, Newcastle University, Newcastle upon Tyne NE1 3BZ, UK.
Rita
R
Horvath
Department of Biosciences, Biotechnology, and Biopharmaceutics, University of Bari Aldo Moro, 70125 Bari, Italy. Electronic address: maria.barile@uniba.it.
Maria
M
Barile
Institute of Human Genetics, Technische Universität München, 81675 Munich, Germany; Institute of Human Genetics, Helmholtz Zentrum München, 85764 Neuherberg, Germany.
Holger
H
Prokisch
D000328:Adult; D015153:Blotting, Western; D016022:Case-Control Studies; D002478:Cells, Cultured; D004579:Electron Transport; D005260:Female; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D005182:Flavin-Adenine Dinucleotide / Q000378:metabolism; D016368:Frameshift Mutation / Q000235:genetics*; D020869:Gene Expression Profiling; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D008099:Liver / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D008297:Male; D028361:Mitochondrial Diseases / Q000188:drug therapy / Q000235:genetics* / Q000473:pathology; D054069:Multiple Acyl Coenzyme A Dehydrogenase Deficiency / Q000188:drug therapy / Q000235:genetics* / Q000473:pathology; D018482:Muscle, Skeletal / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D016297:Mutagenesis, Site-Directed; D009713:Nucleotidyltransferases / Q000235:genetics*; D011485:Protein Binding; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012256:Riboflavin / Q000494:pharmacology*; D012867:Skin / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014803:Vitamin B Complex / Q000494:pharmacology*; D055815:Young Adult
D012333:RNA, Messenger; D014803:Vitamin B Complex; D005182:Flavin-Adenine Dinucleotide; D009713:Nucleotidyltransferases; C020143:FMN adenylyltransferase; D012256:Riboflavin