Sterol uptake induced by an impairment of pyridoxal phosphate synthesis in Saccharomyces cerevisiae: cloning and sequencing of the PDX3 gene encoding pyridoxine (pyridoxamine) phosphate oxidase.
Exogenous sterols do not permeate wild-type Saccharomyces cerevisiae in aerobic conditions. However, mutant strain FKerg7, affected in lanosterol synthase, is a sterol auxotroph which is able to grow aerobically in the presence of ergosterol. Viability of this strain depends on the presence of an additional mutation, aux30, that leads to sterol permeability. Cells bearing the aux30 mutation fail to grow in standard yeast nitrogen base medium containing pyridoxine but grow normally if pyridoxine is replaced by either pyridoxal or pyridoxamine. These mutants are characterized by a lack in pyridoxine (pyridoxamine) phosphate oxidase [P(N/M)P oxidase] (EC 1.4.3.5) activity. The pleiotropic phenotype induced by the aux30 mutation includes a strong perturbation in amino acid biosynthesis. Strains bearing the aux30 mutation also display atypic fatty acid, sterol, and cytochrome patterns. Transformation of an aux30 strain with a replicative vector carrying the wild-type PDX3 gene encoding P(N/M)P oxidase restored wild-type fatty acid, sterol, and cytochrome patterns and suppressed exogenous sterol accumulation. It is proposed that sterol permeation of aux30 strains in mainly the consequence of their leaky Hem- character. The amino acid sequence of S. cerevisiae P(N/M)P oxidase inferred from the nucleotide sequence of PDX3 shows a high percentage of homology with the corresponding enzymes from Escherichia coli and Myxococcus xanthus. Several putative Gcn4p binding sequences are present in the PDX3 promoter region, leading to the assumption that transcription of this gene is under the general control of nitrogen metabolism.
Loubbardi
A
A
Laboratoire de Biochimie et Génétique des Microorganismes, Université de Poitiers, France.
Marcireau
C
C
Karst
F
F
Guilloton
M
M
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA / Q000737:chemistry; D005800:Genes, Fungal; D008969:Molecular Sequence Data; D011732:Pyridoxal Phosphate / Q000096:biosynthesis*; D011734:Pyridoxaminephosphate Oxidase / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D013261:Sterols / Q000378:metabolism*
D013261:Sterols; D011732:Pyridoxal Phosphate; D004247:DNA; D011734:Pyridoxaminephosphate Oxidase