The metG gene of Aspergillus nidulans encoding cystathionine b -lyase: cloning and analysis

Sieńko, Marzena; Paszewski, Andrzej

doi:10.1007/s002940050463

Cited by 24 publications

(19 citation statements)

References 28 publications

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“…The frA gene has been cloned by Ruijter et al (1996); we repeated the "instant gene bank" complementational cloning employed by Ruijter et al and isolated the same genomic sequence, which was used for hybridization with the cosmid library membranes. The cosmid clones containing the methG gene have been identified by Sienko and Paszewski (1999). Finally, J. R. Kinghorn has kindly supplied data on the cosmid clones containing the gdhB gene.…”

Section: Resultsmentioning

confidence: 99%

Genetic and Physical Mapping of Two Centromere-Proximal Regions of Chromosome IV in Aspergillus nidulans

Aleksenko

Nielsen

Clutterbuck

2001

Fungal Genetics and Biology

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Section: Resultsmentioning

confidence: 99%

Genetic and Physical Mapping of Two Centromere-Proximal Regions of Chromosome IV in Aspergillus nidulans

Aleksenko

Nielsen

Clutterbuck

2001

Fungal Genetics and Biology

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“…The two pathways of the trans-sulfuration are, therefore, encoded by dierent sets of genes (Sienko and Paszewski 1999).…”

Section: Discussionmentioning

confidence: 99%

Characterization of the reverse transsulfuration gene mecB of Acremonium chrysogenum, which encodes a functional cystathionine-γ-lyase

et al. 2001

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In Acremonium chrysogenum, biosynthesis of cysteine for the formation of cephalosporin has been proposed to occur through the reverse transsulfuration pathway. A gene, named mecB, has been cloned from an A. chrysogenum C10 genomic library in lambdaEMBL3-ble. The cloned DNA fragment encodes a protein of 423 amino acids with a deduced molecular mass of 45 kDa that shows great similarity to cystathionine-gamma-lyases from Saccharomyces cerevisiae and other eukaryotic organisms. The protein was shown to be functional because it restores growth on methionine to A. nidulans C47 (mecB10), a mutant that is known to be defective in cystathionine-gamma-lyase. The cloned gene did not complement A. nidulans mecA or metG mutants. Enzyme activity assays confirmed that the cloned mecB gene encodes a cystathionine-gamma-lyase activity. The mecB gene is present in a single copy in the wild-type A. chrysogenum (Brotzu's strain) and also in the A. chrysogenum strain C10, a high cephalosporin producer. The gene is localized on chromosome VIII (5.3 Mb), as shown by hybridization to A. chrysogenum chromosomes resolved by pulsed-field gel electrophoresis. Transcription of the mecB gene gives rise to a major transcript of 1.5 kb and a minor one of 1.7 kb. The transcript levels were not significantly affected by addition of DL-methionine to the culture, indicating that expression of this gene is not regulated by methionine. The availability of this gene provides a very useful tool for understanding the proposed role of cystathionine-gamma-lyase in splitting cystathionine to supply cysteine for cephalosporin biosynthesis.

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“…Pfu Turbo (Stratagene) was used for PCRs. The A. nidulans disruption plasmid pJW34 was constructed by ligating a 1.2-kb DNA fragment upstream of the laeA start codon (primers LAE1 and LA2) and a 1.2-kb DNA fragment downstream of the laeA stop codon (primers LA3 and LAE2) to either side of the methG gene in the pUG11-41 vector (31). The 5Ј-end PCR product and 3Ј-end PCR product were inserted into the SacI site and HindIII site of pUG11-41, respectively, by blunt end ligation.…”

Section: Methodsmentioning

confidence: 99%

LaeA, a Regulator of Secondary Metabolism inAspergillusspp

2004

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Secondary metabolites, or biochemical indicators of fungal development, are of intense interest to humankind due to their pharmaceutical and/or toxic properties. We present here a novel Aspergillus nuclear protein, LaeA, as a global regulator of secondary metabolism in this genus. Deletion of laeA (⌬laeA) blocks the expression of metabolic gene clusters, including the sterigmatocystin (carcinogen), penicillin (antibiotic), and lovastatin (antihypercholesterolemic agent) gene clusters. Conversely, overexpression of laeA triggers increased penicillin and lovastatin gene transcription and subsequent product formation. laeA expression is negatively regulated by AflR, a sterigmatocystin Zn 2 Cys 6 transcription factor, in a unique feedback loop, as well as by two signal transduction elements, protein kinase A and RasA. Although these last two proteins also negatively regulate sporulation, ⌬laeA strains show little difference in spore production compared to the wild type, indicating that the primary role of LaeA is to regulate metabolic gene clusters.A complex and fascinating aspect of fungal development is the production of secondary metabolites. These compounds, frequently associated with sporulation processes, are considered part of the chemical arsenal required for niche specialization and have garnered intense interest by virtue of their biotechnological and pharmaceutical applications (9, 11). Many of them display a broad range of useful antibiotic, antiviral, antitumor, antihypercholesterolemic, and immunosuppressant activities as well as less desirable phyto-and mycotoxic activities. Hawksworth's studies of fungal biodiversity led to the conclusion that nearly 1.5 million fungal species exist on Earth, with only 5% identified thus far (16). Thus, the potential for fungal secondary metabolite discovery is vast. Furthermore, the discovery of global regulators for fungal secondary metabolite production is critical, as it would allow for universal manipulation of secondary metabolite production.A large number of known fungal secondary metabolites have been ascribed to the Ascomycete genus Aspergillus. Studies of Aspergillus nidulans have demonstrated the power of using a model system to elucidate the biochemistry and molecular genetics of fungal secondary metabolism, principally penicillin (PN, an antibiotic) and sterigmatocystin (ST, a carcinogen biochemically related to the agricultural contaminant aflatoxin) biosynthesis (6, 17). These studies have established several characteristics of fungal secondary metabolism, including the clustering of biosynthetic and regulatory genes and a genetic connection linking secondary metabolite biosynthesis with sporulation through a shared signal transduction pathway (9).The discovery of the G protein-cyclic AMP (cAMP)-protein kinase A regulation of ST, aflatoxin, and other fungal secondary metabolites (4,17,29,34) has been helpful for establishing a model of global regulation of secondary metabolism. However, all of the signal transduction mutants described in the literature...

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The metG gene of Aspergillus nidulans encoding cystathionine b -lyase: cloning and analysis

Cited by 24 publications

References 28 publications

Genetic and Physical Mapping of Two Centromere-Proximal Regions of Chromosome IV in Aspergillus nidulans

Genetic and Physical Mapping of Two Centromere-Proximal Regions of Chromosome IV in Aspergillus nidulans

Characterization of the reverse transsulfuration gene mecB of Acremonium chrysogenum, which encodes a functional cystathionine-γ-lyase

LaeA, a Regulator of Secondary Metabolism inAspergillusspp

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