title: Comprehensive analysis of expression and function of 51 sarco(endo)plasmic reticulum Ca2+-ATPase mutants associated with Darier disease.
abstract: We examined possible defects of sarco(endo)plasmic reticulum Ca2+-ATPase 2b (SERCA2b) associated with its 51 mutations found in Darier disease (DD) pedigrees, i.e. most of the substitution and deletion mutations of residues reported so far. COS-1 cells were transfected with each of the mutant cDNAs, and the expression and function of the SERCA2b protein was analyzed with microsomes prepared from the cells and compared with those of the wild type. Fifteen mutants showed markedly reduced expression. Among the other 36, 29 mutants exhibited completely abolished or strongly inhibited Ca2+-ATPase activity, whereas the other seven possessed fairly high or normal ATPase activity. In four of the aforementioned seven mutants, Ca2+ transport activity was significantly reduced or almost completely lost, therefore uncoupled from ATP hydrolysis. The other three were exceptional cases as they were seemingly normal in protein expression and Ca2+ transport function, but were found to have abnormalities in the kinetic properties altered by the three mutations, which happened to be in the three DD pedigrees found by us previously (Sato, K., Yamasaki, K., Daiho, T., Miyauchi, Y., Takahashi, H., Ishida-Yamamoto, A., Nakamura, S., Iizuka, H., and Suzuki, H. (2004) J. Biol. Chem. 279, 35595-35603). Collectively, our results indicated that in most cases (48 of 51) DD mutations cause severe disruption of Ca2+ homeostasis by the defects in protein expression and/or transport function and hence DD, but even a slight disturbance of the homeostasis will result in the disease. Our results also provided further insight into the structure-function relationship of SERCAs and revealed critical regions and residues of the enzyme.
authors.affiliation: Departments of Biochemistry and Dermatology, Asahikawa Medical College, Asahikawa 078-8510, Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Miyauchi
authors.forename: Takashi
authors.initials: T
authors.lastname: Daiho
authors.forename: Kazuo
authors.initials: K
authors.lastname: Yamasaki
authors.forename: Hidetoshi
authors.initials: H
authors.lastname: Takahashi
authors.forename: Akemi
authors.initials: A
authors.lastname: Ishida-Yamamoto
authors.forename: Stefania
authors.initials: S
authors.lastname: Danko
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Suzuki
authors.forename: Hajime
authors.initials: H
authors.lastname: Iizuka
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002118:Calcium / Q000737:chemistry / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000235:genetics*; D002522:Chlorocebus aethiops; D007644:Darier Disease / Q000235:genetics*; D007700:Kinetics; D008958:Models, Molecular; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D009154:Mutation; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D013329:Structure-Activity Relationship
chemical_list: D000251:Adenosine Triphosphatases; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000252:Calcium-Transporting ATPases; D002118:Calcium