title: Molecular cloning, characterization and differential expression of Cdc42 in Fonsecaea monophora.
abstract: The cell divisions cycle 42 (Cdc42) gene has been characterized in the fungi, such as Candida albicans, Penicillium marneffei, and Wangiella (Exophiala) dermatitidis, which plays important roles during growth and development. The partial cDNA sequence of Cdc42 of Fonsecaea monophora (F. monophora), designated FmCdc42, was obtained using degenerate primers based on the conserved domain of the other fungi Cdc42. Then the complete cDNA sequence of FmCdc42 was obtained by 5' and 3' RACE. The full-length cDNA is 1,510 bp in size which had an open reading frame (ORF) of 582 bp, encoding 193 amino acid residues. The predicted molecular mass of FmCdc42 is 21.5 kDa with an estimated theoretical isoelectric point of 5.67. The deduced amino acid sequence of FmCdc42 shows 99% identity to that of Wangiella (Exophiala) dermatitidis. 5 exons and 4 introns are identified within the 1,617 bp FmCdc42 genomic DNA sequence of F. monophora. The ORF could be subcloned into the pCDNA6/myc-His B expression vector. The recombinant protein about 27.5 kD infusion protein had high expression level in Vero cells with SDS-PAGE and Western blot analysis. Quantitative real time RT-PCR revealed that FmCdc42 was the highest expression in the sclerotic bodies' stage compared with that in the mycelia and conidia stages, which indicated that the FmCdc42 may be involved in formation of F. monophora sclerotic bodies.
authors.affiliation: Department of Dermatology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, 107 West Yanjiang Road, Guangzhou 510120, China.
authors.forename: Zhi
authors.initials: Z
authors.lastname: Xie
authors.forename: Peiying
authors.initials: P
authors.lastname: Feng
authors.forename: Junming
authors.initials: J
authors.lastname: Zhang
authors.forename: Xiqing
authors.initials: X
authors.lastname: Li
authors.forename: Jiufeng
authors.initials: J
authors.lastname: Sun
authors.forename: Changming
authors.initials: C
authors.lastname: Lu
authors.forename: Huaiqiu
authors.initials: H
authors.lastname: Huang
authors.forename: Liyan
authors.initials: L
authors.lastname: Xi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001203:Ascomycota / Q000235:genetics*; D001483:Base Sequence; D015153:Blotting, Western; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D016000:Cluster Analysis; D019295:Computational Biology; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D016366:Open Reading Frames / Q000235:genetics; D010802:Phylogeny; D060888:Real-Time Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D017385:Sequence Homology; D014709:Vero Cells; D020764:cdc42 GTP-Binding Protein / Q000235:genetics* / Q000378:metabolism*
chemical_list: D017931:DNA Primers; D018076:DNA, Complementary; D020764:cdc42 GTP-Binding Protein