Professor Wlodzimierz Zagorski – biochemist and molecular biologist, focused mainly on studies of structure and physiology of viruses. For many years, as the director of the Institute of Biochemistry and Biophysics of the Polish Academy of Sciences he established a close collaboration between the Institute and several Polish and foreign research institutions. He promoted the integration of Polish research teams with international research community by their participation in multinational programs and consortia. He contributed much to the development of biotechnology in Poland and to creating links between basic research and economy. Besides being a scientist he was well known as intellectualist attaching a great importance to cultural, social and political matters of the state.

In Aspergillus nidulans sulfate is taken up by sulfate permease encoded by the sB gene. A unique tight auxotrophic mutant with an impaired promoter region of the sulfate permease gene, sB1pr, was isolated. Three suppressor genes were cloned by complementation of this mutation. One of them, described here, is the astA gene (alternative sulfate transporter) derived from a genomic library of the Japanese A. nidulans IAM 2006 strain. In the reference strain of Glasgow origin the astA gene was found to be a pseudogene having several nucleotide deletions in ORF. The gene encodes a novel type of sulfate transporter which is distinct from other known sulfate permeases forming the SulP family. The putative ASTA protein belongs to an extensive and poorly characterized Dal5 allantoate permease family of fungal organic anion transporters. We have shown that ASTA is a physiological sulfate transporter. We also report cloning and characterization of the sB gene in this work. Both genes, sB and astA, are regulated at the transcriptional level by sulfur metabolite repression (SMR).

The identification, isolation and characterization of a new Aspergillus nidulans positive-acting gene metR, which encodes a transcriptional activator of sulphur metabolism, is reported. metR mutants are tight auxotrophs requiring methionine or homocysteine for growth. Mutations in the metR gene are epistatic to mutations in the negative-acting sulphur regulatory scon genes. The metR coding sequence is interrupted by a single intron of 492 bp which is unusually long for fungi. Aspergillus nidulans METR is a member of bZIP family of DNA-binding proteins. The bZIP domains of METR and the Neurospora crassa CYS3 transcriptional activator of sulphur genes are highly similar. Although Neurospora cys-3 gene does not substitute for the metR function, a chimeric metR gene with a cys-3 bZIP domain is able to transform the DeltametR mutant to methionine prototrophy. This indicates that METR recognizes the same regulatory sequence as CYS3. The metR gene is not essential, as deletion mutants are viable and have similar phenotype as point mutants. In contrast to the Neurospora cys-3, transcription of the metR gene was found to be regulated neither by METR protein nor by sulphur source. Transcription of metR gene is derepressed in the sconB2 mutant. Transcription of genes encoding sulphate permease, homocysteine synthase, cysteine synthase, ATP-sulphurylase, and sulphur controller-sconB is strongly regulated by the metR gene product and depends on the character of the metR mutation and sulphur supplementation.

Roles played by homocysteine and choline in the regulation of MS (methionine synthase) have been examined in fungi. The Aspergillus nidulans metH gene encoding MS was cloned and characterized. Its transcription was not regulated by methionine, but was enhanced by homocysteine and repressed by choline and betaine. MS activity levels were regulated in a similar way. The repression by betaine was due to its metabolic conversion to choline, which was found to be very efficient in A. nidulans. Betaine and choline supplementation stimulated growth of leaky metH mutants apparently by decreasing the demand for methyl groups and thus saving methionine and S-adenosylmethionine. We have also found that homocysteine stimulates transcription of MS-encoding genes in Saccharomyces cerevisiae and Schizosaccharomyces pombe.

Mutations in the Aspergillus nidulans metE gene lead to requirement for O-acetylhomoserine. The gene was cloned by complementation of the metE31 mutation. The coding sequence was found to be interrupted by two introns of 66 and 50 bp, respectively. metE codes for a peptide of 489 amino acids which belongs to the family of homoserine O-acetyltransferases and a well-defined superfamily of alpha/beta hydrolases. Transcription of the metE gene is strongly up-regulated by a severe limitation of methionine, but not of cysteine. This gene is the first sulphur metabolism gene described in A. nidulans which is not regulated by the sulphur metabolite repression system in which cysteine acts as the low-molecular-weight effector.

Conference Abstract| October 01 2000 Pleiotropic effects of mutation in the methionine synthase gene in Aspergillus nidulans M. Kacprzak; M. Kacprzak 1Institute of Biochemistry and Biophysics, ul. Pawinskiego 5A, Warsaw Poland Search for other works by this author on: This Site PubMed Google Scholar I. Lewandowska; I. Lewandowska 1Institute of Biochemistry and Biophysics, ul. Pawinskiego 5A, Warsaw Poland Search for other works by this author on: This Site PubMed Google Scholar M. Balinska; M. Balinska 2Institute of Experimental Biology, ul. Pasteura 3. Warsaw. Poland Search for other works by this author on: This Site PubMed Google Scholar A. Paszewski A. Paszewski 1Institute of Biochemistry and Biophysics, ul. Pawinskiego 5A, Warsaw Poland Search for other works by this author on: This Site PubMed Google Scholar Biochem Soc Trans (2000) 28 (5): A194. https://doi.org/10.1042/bst028a194 Views Icon Views Article contents Figures & tables Video Audio Supplementary Data Peer Review Share Icon Share Facebook Twitter LinkedIn MailTo Cite Icon Cite Get Permissions Citation M. Kacprzak, I. Lewandowska, M. Balinska, A. Paszewski; Pleiotropic effects of mutation in the methionine synthase gene in Aspergillus nidulans. Biochem Soc Trans 1 October 2000; 28 (5): A194. doi: https://doi.org/10.1042/bst028a194 Download citation file: Ris (Zotero) Reference Manager EasyBib Bookends Mendeley Papers EndNote RefWorks BibTex toolbar search Search Dropdown Menu toolbar search search input Search input auto suggest filter your search All ContentAll JournalsBiochemical Society Transactions Search Advanced Search This content is only available as a PDF. © 2000 Biochemical Society2000 Article PDF first page preview Close Modal You do not currently have access to this content.

sconC, which encodes a negative regulator of sulphur metabolism in Aspergillus nidulans was cloned, sequenced, and found to belong to the highly conserved family of SKP1 genes essential for many cell functions, including cell cycle regulation. The ORF of 722 bp, encoding a protein of 161 amino acids, is interrupted by four introns. There is a fifth intron (135 bp long) in the upstream untranslated sequence. Two point mutations in conserved regions were identified in the mutant alleles sconC3 and sconC1, which result in relief of sulphur metabolite repression. The SCONC protein contains the PEST sequence common for proteins that are subject to rapid turnover. Transformation of the sconC3 mutant with sconB+ restores the wild-type phenotype. The sconB gene encodes a protein containing the F-box, a domain known to interact with Skp1 proteins. By analogy with other systems, it seems likely that the SCONC protein interacts with SCONB. sconC mRNA is present in the sconC3 and sconB2 mutants and the level of the sconC transcript seems not to be significantly regulated by supplementation of the medium with sulphur.

The Aspergillus nidulans cysA gene was cloned by functional complementation of the cysA1 mutation that impairs the synthesis of O:-acetylserine. The molecular nature of cysA1 and cysA103 alleles was characterized; a nucleotide substitution and a frame shift were found in the former and a deletion mutation in the latter. The CYSA protein is 525 amino acids long and is encoded by an uninterrupted open reading frame. Expression of the cysA gene appears not to be regulated by sulfur, carbon and nitrogen sources. Protein sequence analysis reveals extensive similarity to homoserine O:-acetyltransferases, particularly the bacterial ones, and no homology with known serine O:-acetyltransferases. The authors propose that the CYSA protein is analogous to serine O:-acetyltransferases, i.e. it catalyses the same reaction but has an independent evolutionary origin.

Using a Saccharomyces cerevisiae strain having the activities of serine O-acetyl-transferase (SATase), O-acetylserine/O-acetylhomoserine sulphydrylase (OAS/OAH SHLase), cystathionine β-synthase (β-CTSase) and cystathionine γ-lyase (γ-CTLase), we individually disrupted CYS3(coding for γ-CTLase) and CYS4 (coding for β-CTSase). The obtained gene disruptants were cysteine-dependent and incorporated the radioactivity of 35S-sulphate into homocysteine but not into cysteine or glutathione. We concluded, therefore, that SATase and OAS/OAH SHLase do not constitute a cysteine biosynthetic pathway and that cysteine is synthesized exclusively through the pathway constituted with β-CTSase and γ-CTLase; note that OAS/OAH SHLase supplies homocysteine to this pathway by acting as OAH SHLase. From further investigation upon the cys3-disruptant, we obtained results consistent with our earlier suggestion that cysteine and OAS play central roles in the regulation of sulphate assimilation. In addition, we found that sulphate transport activity was not induced at all in the cys4-disruptant, suggesting that CYS4 plays a role in the regulation of sulphate assimilation. Copyright © 1999 John Wiley & Sons, Ltd.

The metG gene of Aspergillus nidulans encoding cystathionine β-lyase, an enzyme of the main pathway of methionine synthesis, was cloned by complementation of a metG mutation. A comparison of metG genomic DNA and a cDNA copy derived from different A. nidulans strains revealed a marked DNA sequence polymorphism manifested mostly by silent point mutations. cDNA of the A. nidulans metG gene complemented the Escherichia coli metC69 mutation impairing cystathionine β-lyase. This gene contains two introns and codes for a protein of 439 amino acids. The protein shows homology with bacterial, yeast and plant cystathionine β-lyases, as well as with other enzymes belonging to a large family of pyridoxal 5′-phosphate binding proteins. Transcription of the metG gene is not appreciably regulated by the concentration of sulphur source in the growth medium.

The Kluyveromyces lactis homocysteine synthase gene was cloned by complementation of the Saccharomyces cerevisiae met25 mutation. The coding sequence of the K. lactis gene shows a high similarity to the S. cerevisiae gene. Very little similarity is found in the 5' and 3' untranslated regions. However, one finds short DNA stretches in the promoter of the K. lactis gene which are identical to the nucleotide sequences implicated in the regulation of the S. cerevisiae homologue. This could explain strong transcriptional inhibition of the K. lactis gene by exogenous methionine in the S. cerevisiae host, and indicates a substantial conservation of the sulphur regulatory system between both yeast species.