title: Novel carboxypeptidase A6 (CPA6) mutations identified in patients with juvenile myoclonic and generalized epilepsy.
abstract: Carboxypeptidase A6 (CPA6) is a peptidase that removes C-terminal hydrophobic amino acids from peptides and proteins. The CPA6 gene is expressed in the brains of humans and animals, with high levels of expression during development. It is translated with a prodomain (as proCPA6), which is removed before secretion. The active form of CPA6 binds tightly to the extracellular matrix (ECM) where it is thought to function in the processing of peptides and proteins. Mutations in the CPA6 gene have been identified in patients with temporal lobe epilepsy and febrile seizures. In the present study, we screened for CPA6 mutations in patients with juvenile myoclonic epilepsy and identified two novel missense mutations: Arg36His and Asn271Ser. Patients harboring these mutations also presented with generalized epilepsy. Neither of the novel mutations was found in a control population. Asn271 is highly conserved in CPA6 and other related metallocarboxypeptidases. Arg36 is present in the prodomain and is not highly conserved. To assess structural consequences of the amino acid substitutions, both mutants were modeled within the predicted structure of the enzyme. To examine the effects of these mutations on enzyme expression and activity, we expressed the mutated enzymes in human embryonic kidney 293T cells. These analyses revealed that Asn271Ser abolished enzymatic activity, while Arg36His led to a ~50% reduction in CPA6 levels in the ECM. Pulse-chase using radio-labeled amino acids was performed to follow secretion. Newly-synthesized CPA6 appeared in the ECM with peak levels between 2-8 hours. There was no major difference in time course between wild-type and mutant forms, although the amount of radiolabeled CPA6 in the ECM was lower for the mutants. Our experiments demonstrate that these mutations in CPA6 are deleterious and provide further evidence for the involvement of CPA6 mutations in the predisposition for several types of epilepsy.
authors: Department of Neuroscience, Albert Einstein College of Medicine, Bronx, NY, United States of America.
authors: Matthew R
authors: MR
authors: Sapio
authors: Department of Genetic Medicine and Laboratory, University Hospitals of Geneva, Geneva, Switzerland.
authors: Monique
authors: M
authors: Vessaz
authors: Department of Neurology, University Hospital, Nice, France.
authors: Pierre
authors: P
authors: Thomas
authors: Centre Saint Paul, Hôpital Henri Gastaut, Marseille, France.
authors: Pierre
authors: P
authors: Genton
authors: Department of Neuroscience, Albert Einstein College of Medicine, Bronx, NY, United States of America; Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, NY, United States of America.
authors: Lloyd D
authors: LD
authors: Fricker
authors: Department of Psychiatry, University of Geneva, Geneva, Switzerland.
authors: Annick
authors: A
authors: Salzmann
mesh_terms: D000293:Adolescent; D000328:Adult; D000483:Alleles; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000927:Anticonvulsants / Q000627:therapeutic use; D043422:Carboxypeptidases A / Q000737:chemistry / Q000235:genetics*; D016022:Case-Control Studies; D002648:Child; D002675:Child, Preschool; D004252:DNA Mutational Analysis; D004829:Epilepsy, Generalized / Q000175:diagnosis / Q000188:drug therapy / Q000235:genetics*; D005260:Female; D015870:Gene Expression; D056726:Genetic Association Studies; D020022:Genetic Predisposition to Disease; D006801:Humans; D008297:Male; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D020190:Myoclonic Epilepsy, Juvenile / Q000175:diagnosis / Q000188:drug therapy / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide; D011487:Protein Conformation; D016415:Sequence Alignment; D055815:Young Adult
chemical_list: D000927:Anticonvulsants; C578288:CPA6 protein, human; D043422:Carboxypeptidases A