Cloning and expression of the ARO3 gene encoding DAHP synthase from Candida albicans

Pereira, Sarita A.; Livi, George P.

doi:10.1016/0378-1119(93)90191-5

Cited by 14 publications

(13 citation statements)

References 32 publications

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“…As shown in Fig. 4(a, b), the C. albicans ARO4 gene, like ARO3 (Pereira & Livi, 1993), complements the aro3 aro4 mutations in S. cerevisiae. The ARO3 gene from C. albicans was previously defined by its inability to complement when cells are grown on excess (2 mM) phenylalanine, presumably due to isozyme-specific feedback inhibition (Pereira & Livi, 1993).…”

Section: Functional Expression In S Cerevisiaementioning

confidence: 72%

“…4(a, b), the C. albicans ARO4 gene, like ARO3 (Pereira & Livi, 1993), complements the aro3 aro4 mutations in S. cerevisiae. The ARO3 gene from C. albicans was previously defined by its inability to complement when cells are grown on excess (2 mM) phenylalanine, presumably due to isozyme-specific feedback inhibition (Pereira & Livi, 1993). Complemen- (Pereira & Livi, 1993) or the C. albicans ARO4 gene were introduced into cells of S. cerevisiae strain RH1368 (aro3-2 aro4-1 gcn1-1 trp1-1 leu2-3), selecting for Leu + transformants.…”

Section: Functional Expression In S Cerevisiaementioning

confidence: 97%

“…We previously generated a 222 bp genomic PCR fragment from a homozygous aro3-deletion strain of C. albicans, using a degenerate primer based on a conserved N-terminal region of Aro3p (Pereira & Livi, 1993), plus a degenerate comeback primer encoding a conserved region of the protein that lies within the deleted portion of the gene (Pereira & Livi, 1996). The nucleotide sequence of this PCR fragment predicts a 74 aa DAHPsynthase-related peptide with strong homology to known Aro3p\Aro4p proteins.…”

Section: Cloning and Characterization Of The C Albicans Aro4 Genementioning

confidence: 99%

“…The ARO3 gene from C. albicans was previously defined by its inability to complement when cells are grown on excess (2 mM) phenylalanine, presumably due to isozyme-specific feedback inhibition (Pereira & Livi, 1993). Complemen- (Pereira & Livi, 1993) or the C. albicans ARO4 gene were introduced into cells of S. cerevisiae strain RH1368 (aro3-2 aro4-1 gcn1-1 trp1-1 leu2-3), selecting for Leu + transformants. Numbered spots correspond to the following strains in each panel : spot 1, RH1368 plus YEp13-ARO3 ; spot 2, RH1368 plus YEp13-ARO4 ; spot 3, RH1368 plus YEp13 alone ; spot 4, untransformed RH1368 ; spot 5, untransformed strain DC17 (MATα his1 ; Klar et al, 1985).…”

Section: Functional Expression In S Cerevisiaementioning

confidence: 99%

“…We are studying the nature and complexity of DAHP synthases in the pathogenic fungi as potential antifungal drug targets, since : (1) humans lack a comparable biosynthetic pathway -instead they rely on dietary sources for phenylalanine and tryptophan, and can only synthesize tyrosine via hydroxylation of phenylalanine ; (2) based on pathogenicity studies of other auxotrophic mutants of Candida albicans and Cryptococcus neoformans (Manning et al, 1984 ;Shepherd, 1985 ;Kirsch & Whitney, 1991 ;Perfect et al, 1993), aromatic amino acid auxotrophs are predicted to display decreased virulence in vivo as a result of poor growth due to suboptimal amino acid bioavailability ; and (3) certain amino acid biosynthesis inhibitors have been used safely and effectively as herbicides (reviewed by Kishore & Shah, 1988). We previously cloned an ARO3 gene orthologue from the diploid pathogenic fungus C. albicans and found that it can complement an aro3 aro4 double mutation in S. cerevisiae, and that complementation is inhibited by excess phenylalanine (Pereira & Livi, 1993). Expression of C. albicans ARO3 mRNA is induced in response to amino acid starvation, consistent with the presence of two putative GCREs in the promoter sequence (Pereira & Livi, 1995).…”

Section: Introductionmentioning

confidence: 99%

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The ARO4 gene of Candida albicans encodes a tyrosine-sensitive DAHP synthase: evolution, functional conservation and phenotype of Aro3p-, Aro4p-deficient mutants The GenBank accession number for the sequence reported in this paper is U53216.

et al. 2002

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The enzyme 3-deoxy-D-arabinoheptulosonate-7-phosphate (DAHP) synthase catalyses the first step in aromatic amino acid biosynthesis in prokaryotes, plants and fungi. Cells of Saccharomyces cerevisiae contain two catalytically redundant DAHP synthases, encoded by the genes ARO3 and ARO4, whose activities are feedback-inhibited by phenylalanine and tyrosine, respectively. ARO3/4 gene transcription is controlled by GCN4. The authors previously cloned an ARO3 gene orthologue from Candida albicans and found that : (1) it can complement an aro3 aro4 double mutation in S. cerevisiae, an effect inhibited by excess phenylalanine, and (2) a homozygous aro3-deletion mutant of C. albicans is phenotypically Aro M , suggesting the existence of another isozyme(s). They now report the identification and functional characterization of the C. albicans orthologue of S. cerevisiae Aro4p. The two Aro4p enzymes share 68 % amino acid identity. Phylogenetic analysis places the fungal DAHP synthases in a cluster separate from prokaryotic orthologues and suggests that ARO3 and ARO4 arose from a single gene via a gene duplication event early in fungal evolution. C. albicans ARO4 mRNA is elevated upon amino acid starvation, consistent with the presence of three putative Gcn4p-responsive elements (GCREs) in the gene promoter sequence. C. albicans ARO4 complements an aro3 aro4 double mutation in S. cerevisiae, an effect inhibited by excess tyrosine. The authors engineered ∆aro3/∆aro3 ∆aro4/MET3p ::ARO4 cells of C. albicans (with one wild-type copy of ARO4 placed under control of the repressible MET3 promoter) and found that they fail to grow in the absence of aromatic amino acids when ARO4 expression is repressed, and that this growth defect can be partially rescued by aromatic amino acids and certain aromatic amino acid pathway intermediates. It is concluded that, like S. cerevisiae, C. albicans contains two DAHP synthases required for the first step in the aromatic amino acid biosynthetic pathway.

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Section: Functional Expression In S Cerevisiaementioning

confidence: 72%