title: Roles of phytanoyl-CoA alpha-hydroxylase in mediating the expression of human coagulation factor VIII.
abstract: The coagulation factor VIII (FVIII) is the coagulation factor deficient in the X-chromosome-linked bleeding disorder hemophilia A. Previous transfection studies demonstrated that factor VIII was 10-100-fold less efficiently expressed than the homologous coagulation factor, factor V. To investigate the regulatory mechanisms of FVIII synthesis and secretion, we used the yeast two-hybrid system as an approach to search for proteins that associated with FVIII. The A2 domain (337-740 amino acids) of factor VIII (FVIII-A2) was used as a bait and phytanoyl-CoA alpha-hydroxylase (PAHX) was identified as a binding protein of FVIII-A2. PAHX had potential to interact with the residues 373-508 within the A2 domain, but not with A1 and A3 (the homologous domains of A2). The interaction between the A2 domain and PAHX was independent of the type 2 peroxisomal targeting signal (PTS2) of PAHX. Overexpression of PAHX in FVIII-produced cells decreased the expression of FVIII by about 70%. The elevated expression of von Willebrand factor had no effect on the suppression of FVIII secretion by PAHX. Expression of the green fluorescent PAHX fusion protein in SMMC-7721 cells affected the intracellular trafficking of FVIII-A2. These results suggested that the interaction between PAHX and FVIII-A2 was in part responsible for the low-level expression of factor VIII.
authors.affiliation: Gene Research Center, Medical Center of Fudan University, Shanghai 200032, China.
authors.forename: C
authors.initials: C
authors.lastname: Chen
authors.forename: Q
authors.initials: Q
authors.lastname: Wang
authors.forename: X
authors.initials: X
authors.lastname: Fang
authors.forename: Q
authors.initials: Q
authors.lastname: Xu
authors.forename: C
authors.initials: C
authors.lastname: Chi
authors.forename: J
authors.initials: J
authors.lastname: Gu
mesh_terms: D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D003001:Cloning, Molecular; D006224:Cricetinae; D017931:DNA Primers; D005169:Factor VIII / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation / Q000502:physiology*; D006801:Humans; D006899:Mixed Function Oxygenases / Q000378:metabolism / Q000502:physiology*; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D017931:DNA Primers; D011993:Recombinant Fusion Proteins; D005169:Factor VIII; D006899:Mixed Function Oxygenases; C102623:PHYH protein, human