Summary
The structure, function and regulation of the acetate inducible aciA gene of Aspergillus nidulans was analysed. The aciA locus was mapped to chromosome 1 at a position where no acetate inducible gene has been previously located. The nucleotide sequence of aciA was determined, the structures of two transcripts were determined and the sequences of the polypeptide products of the gene were deduced. Construction of an aciA loss-of-function mutant was achieved via insertional inactivation, but it did not reveal a phenotype for an aciA − strain. The larger polypeptide, AciA, was found to have a putative dinucleotide cofactor binding site. Acetate inducibility of aciA was found to be dependent on the amdA regulatory gene. Use of a 5′ deletion series of an aciA-lacZ fusion and an in vivo regulatory protein binding (titration) assay allowed the region required for amdA activity to be localized to a 124 by segment 5′ to aciA. A 13 by region of sequence similarity was observed between aciA and the coregulated amdS gene in the regions required for amdA-mediated regulation of these genes. This sequence may have a role in amdA regulation of the two genes.
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References
Andrianopoulos A, Hynes MJ (1988) Cloning and analysis of the positively acting regulatory gene amdR from Aspergillus nidulans. Mol Cell Biol 8:3532–3541
Armitt S, McCullough W, Roberts CF (1976) Analysis of acetate nonutilising (acu) mutants in Aspergillus nidulans. J Gen Microbiol 92:263–282
Arst HN (1976) Integrator gene in Aspergillus nidulans. Nature 262:231–234
Atkinson PW, King JA, Hynes MJ (1985) Identification of the aciA gene controlled by the amdA regulatory gene in Aspergillus nidulans. Curr Genet 10:133–138
Ballance DJ (1986) Sequences important for gene expression in filamentous fungi. Yeast 2:229–236
Burke JF (1984) Highly-sensitive S1 mapping with single-stranded [32P] DNA probes synthesized from bacteriophage M13mp9 templates. Gene 30:63–68
Casadaban MJ, Martinez-Arias A, Shapira SK, Chou J (1983) Galactosidase gene fusions for analysing gene expression in Escherichia coli and yeast. Methods Enzymol 100:293–307
Chou PY, Fasman GD (1974) Prediction of protein conformation. Biochemistry 13:222–245
Clutterbuck AJ (1974) Aspergillus nidulans genetics. In: King RC (ed) Handbook of genetics, vol 1. Plenum Press, New York
Corpet F (1988) Multiple sequence alignment with hierarchical clustering. Nucleic Acids Res 16:10881–10890
Corrick CM, Twomey AP, Hynes MJ (1987) The nucleotide sequence of the amdS gene of Aspergillus nidulans and the molecular characterization of 5′ mutations. Gene 53:63–71
Cove DJ (1966) The induction and repression of nitrate reductase in the fungus Aspergillus nidulans. Biochem Biophys Acta 113:51–56
Dale RM, McClure Ba, Houchins JP (1985) A rapid singlestranded cloning strategy for producing a sequential series of overlapping clones for use in DNA sequencing: application to sequencing the corn mitochondrial 18 S rDNA. Plasmid 13:31–40
Davis MA, Cobbett CS, Hynes MJ (1988) An amdS-lacZ fusion for studying gene regulation in Aspergillus. Gene 63:199–212
Durrens P, Green PM, Arst HN Jr, Scazzocchio C (1986) Heterologous insertion of transforming DNA and generation of new deletions associated with transformation in Aspergillus nidulans. Mol Gen Genet 203:544–549
Greenler JM, Sloan JS, Schwartz BW, Becker WM (1989) Isolation, characterization and sequence analysis of a full-length cDNA clone encoding NADH-dependent hydroxypyruvate reductase from cucumber. Plant Mol Biol 13:139–150
Gurr SJ, Unkles SE, Kinghorn JR (1987) The structure and organisation of nuclear genes of filamentous fungi. In: Kinghom JR (ed) Gene Structure in Eukaryotic Microbes. IRL press, Oxford, pp 93–139
Hynes MJ (1977) Induction of the acetamidase of Aspergillus nidulans by acetate metabolism. J Bacteriol 131:770–775
Hynes MJ (1978) Multiple independent control mechanisms affecting the acetamidase of Aspergillus nidulans. Mol Gen Genet 161:59–65
Hynes MJ (1982) A cis-dominant mutation in Aspergillus nidulans affecting the expression of the amdS gene in the presence of mutations in the unlinked gene amdA. Genetics 102:139–147
Hynes MJ, Pateman JA (1970a) The genetic analysis of regulation of amidase synthesis in Aspergillus nidulans. I. Mutants able to use acrylamide. Mol Gen Genet 108:95–106
Hynes MJ, Pateman JA (1970b) The genetic analysis of regulation of amidase synthesis in Aspergillus nidulans. II. Mutants resistant to fluoroacetamide. Mol Gen Genet 108:107–116
Hynes MJ, Corrick CM, King JA (1983) Isolation of genomic clones containing the amdS gene of Aspergillus nidulans and their use in the analysis of structural and regulatory mutations. Mol Cell Biol 3:1430–1439
Hynes MJ, Corrick CM, Kelly JM, Littlejohn TG (1988) Identification of the sites of action for regulatory genes controlling the amdS gene of Aspergillus nidulans. Mol Cell Biol 8:2589–2596
Hynes MJ, Andrianopoulos A, Davis MA, van Heejswick R, Katz ME, Littlejohn TG, Richardson IB, Saleeba JA (1989) Multiple circuits regulating the amdS gene of Aspergillus nidulans. In: Penttila M, Nevalainen H (eds) Proceedings of the EMBO-Alko Workshop on Molecular Biology of Filamentous Fungi. Foundation for Biotechnical and Industrial Fermentation Research 6, Helsinki, pp 63–71
Katz ME, Saleeba JA, Sapats SI, Hynes MJ (1990) A mutation affecting amdS expression in Aspergillus nidulans contains a triplication of a cis-acting regulatory sequence. Mol Gen Genet 220:373–376
Lerch H-P, Blocker H, Kallwass H, Hoppe J, Tsai H, Collins J (1989) Cloning, sequencing and expression in Escherichia coli of the D-2-hydroxyisocaproate dehydrogenase gene of Lactobacillus casei. Gene 78:47–57
Lipman DJ, Pearson WR (1985) Rapid and sensitive protein similarity searches. Science 227:1435–1441
Messing J, Crea R, Seeburg PH (1981) A system for shotgun DNA sequencing. Nucleic Acids Res 9:309–323
Norrander J, Kempe T, Messing J (1983) Construction of improved M13 vectors using oligodeoxynucleotide-directed mutagenesis. Gene 26:101–106
Pearson WR, Lipman DJ (1988) Improved tools for biological sequence comparison. Proc Natl Acad Sci USA 85:2444–2448
Reed KC, Mann DA (1985) Rapid transfer of DNA from agarose gels to nylon membranes. Nucleic Acids Res 13:7207–7221
Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York
Sanger F, Nicklen S, Coulson AR (1977) DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci USA 74:5463–5467
Schoenlein PV, Rea BB, Winkler ME (1989) Divergent transcription of pdxB and homology between the pdxB and serA gene products in Escherichia coli K-12. J Bacteriol 171:6084–6092
Southern EM (1975) Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol 98:503–517
Tobey KL, Grant GA (1986) The nucleotide sequence of the serA gene of Escherichia coli and the amino acid sequence of the encoded protein, D-3-phosphoglycerate dehydrogenase. J Biol Chem 261:12179–12183
Wierenga RK, Terpstra P, Hol WGJ (1986) Prediction of the occurence of the ADP-binding βαβ-fold in proteins, using an amino acid sequence fingerprint. J Mol Biol 187:101–107
Yelton MM, Hamer JE, Timberlake WE (1984) Transformation of Aspergillus nidulans by using a trpC plasmid. Proc Natl Acad Sci USA 81:1470–1474
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Communicated by C.A.M.J.J. van den Hondel
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Saleeba, J.A., Cobbett, C.S. & Hynes, M.J. Characterization of the amdA-regulated aciA gene of Aspergillus nidulans . Molec. Gen. Genet. 235, 349–358 (1992). https://doi.org/10.1007/BF00279380
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DOI: https://doi.org/10.1007/BF00279380