title: Identification and characterization of two novel mutations (Q421 K and R123P) in congenital factor XII deficiency.
abstract: The factor XII genes of two unrelated factor XII-deficient Japanese families were screened, and two novel mutations were identified. A heterozygous mutation (Q421K) was identified in the gene of a cross-reacting material (CRM)-negative patient with reduced FXII activity (entitled Case 1). No mutations were discovered in the other allele. Case 2 was a CRM-negative patient with severe FXII deficiency. In this case, a homozygous mutation (R123P) was discerned. Expression studies in Chinese Hamster Ovary (CHO) cells demonstrated accumulation of mutant Q421 K factor XII in the cell, and insufficient secretion, while the R123P mutant showed lower levels of accumulation than wild-type, and no evidence of secretion in culture supernatant. In the presence of proteasome inhibitor, all types of FXII (wild-type. Q421K, R123P) accumulated in the cells. Protease protection experiments using the microsomal fraction of these cell lines demonstrated that while 20% wild-type FXII (total wild-type:100%) and 10% R123P mutant (total R123P-type: 40%) were resistant to treatment with trypsin, 50% Q421K-type FXII (total Q421K-type:130%) remained resistant to digestion. From these results, we conclude that Q421K is less susceptible to proteasome degradation than wild-type, but is unable to exit the ER efficiently, resulting in insufficient secretion phenotype. In contrast, R123P is susceptible to proteasome degradation and is not secreted.
authors.affiliation: Kyushu University Graduate School of Medicine, The First Department of Internal Medicine, Faculty of Medicine, Kyushu University, Fukuoka, Japan. tkana@scripps.edu
authors.forename: T
authors.initials: T
authors.lastname: Kanaji
authors.forename: S
authors.initials: S
authors.lastname: Kanaji
authors.forename: K
authors.initials: K
authors.lastname: Osaki
authors.forename: M
authors.initials: M
authors.lastname: Kuroiwa
authors.forename: M
authors.initials: M
authors.lastname: Sakaguchi
authors.forename: K
authors.initials: K
authors.lastname: Mihara
authors.forename: Y
authors.initials: Y
authors.lastname: Niho
authors.forename: T
authors.initials: T
authors.lastname: Okamura
mesh_terms: D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000293:Adolescent; D019943:Amino Acid Substitution; D000818:Animals; D020126:Brefeldin A / Q000494:pharmacology; D016466:CHO Cells; D003062:Codon / Q000235:genetics; D006224:Cricetinae; D003412:Cricetulus; D003546:Cysteine Endopeptidases / Q000378:metabolism; D004252:DNA Mutational Analysis; D005091:Exons / Q000235:genetics; D005174:Factor XII / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D005175:Factor XII Deficiency / Q000235:genetics*; D006579:Heterozygote; D006720:Homozygote; D006801:Humans; D008297:Male; D008875:Middle Aged; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors / Q000378:metabolism; D020125:Mutation, Missense; D010314:Partial Thromboplastin Time; D010375:Pedigree; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D011480:Protease Inhibitors / Q000494:pharmacology; D046988:Proteasome Endopeptidase Complex; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection
chemical_list: D003062:Codon; D009097:Multienzyme Complexes; D011480:Protease Inhibitors; D011993:Recombinant Fusion Proteins; C067713:lactacystin; D020126:Brefeldin A; D005174:Factor XII; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D000111:Acetylcysteine