Isolation and analysis of functional homologues of the secretion-related SAR1 gene of Saccharomyces cerevisiae from Aspergillus niger and Trichoderma reesei

G. Veldhuisen, M. Saloheimo, M. A. Fiers, P. J. Punt, R. Contreras, M. Penttilä, C. A.M.J.J. Van Den Hondel

Research output: Contribution to journalArticleScientificpeer-review

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Abstract

The Aspergillus niger and Trichoderma reesei genes encoding the functional homologues of the small GTP-binding protein SAR1p, which is involved in the secretion pathway in Saccharomyces cerevisiae, have been cloned and characterised. The A. niger gene (sarA) contains five introns, whereas the T. reesei gene (sar1) has only four. In both cases the first intron is at the same position as the single S. cerevisiae SAR1 intron. The encoded proteins show 70-80% identity to the SAR1 protein. Complementation of S. cerevisiae sar1 and sec12 mutants by expression vectors carrying the A. niger sarA and T. reesei sar1 cDNA clones confirmed that the cloned genes are functional homologues of the S. cerevisiae SAR1 gene. Three mutant alleles of the A. niger sarA gene (D29G, E109K, D29G/E109K), generated by site-directed mutagenesis, revealed a thermosensitive dominant-negative phenotype in the presence of the wild-type sarA allele. This result contrasts with the situation in S. cerevisiae, where similar mutations have a thermosensitive phenotype. Taken together, our results indicate that the sarA gene is involved in an essential function in A. niger.

Original languageEnglish
Pages (from-to)446-455
Number of pages10
JournalMolecular and General Genetics
Volume256
Issue number4
DOIs
Publication statusPublished - 27 Nov 1997
MoE publication typeA1 Journal article-refereed

Fingerprint

Trichoderma
Aspergillus niger
Saccharomyces cerevisiae
Genes
Introns
Alleles
Phenotype
Secretory Pathway
Site-Directed Mutagenesis
GTP-Binding Proteins
Proteins
Complementary DNA
Clone Cells
Mutation

Keywords

  • Complementation
  • Filamentous fungi
  • Gene cloning
  • Protein secretion
  • Small GTP binding protein

Cite this

@article{9ccf60f8ef32463e8e0b4c65d2ca66bd,
title = "Isolation and analysis of functional homologues of the secretion-related SAR1 gene of Saccharomyces cerevisiae from Aspergillus niger and Trichoderma reesei",
abstract = "The Aspergillus niger and Trichoderma reesei genes encoding the functional homologues of the small GTP-binding protein SAR1p, which is involved in the secretion pathway in Saccharomyces cerevisiae, have been cloned and characterised. The A. niger gene (sarA) contains five introns, whereas the T. reesei gene (sar1) has only four. In both cases the first intron is at the same position as the single S. cerevisiae SAR1 intron. The encoded proteins show 70-80{\%} identity to the SAR1 protein. Complementation of S. cerevisiae sar1 and sec12 mutants by expression vectors carrying the A. niger sarA and T. reesei sar1 cDNA clones confirmed that the cloned genes are functional homologues of the S. cerevisiae SAR1 gene. Three mutant alleles of the A. niger sarA gene (D29G, E109K, D29G/E109K), generated by site-directed mutagenesis, revealed a thermosensitive dominant-negative phenotype in the presence of the wild-type sarA allele. This result contrasts with the situation in S. cerevisiae, where similar mutations have a thermosensitive phenotype. Taken together, our results indicate that the sarA gene is involved in an essential function in A. niger.",
keywords = "Complementation, Filamentous fungi, Gene cloning, Protein secretion, Small GTP binding protein",
author = "G. Veldhuisen and M. Saloheimo and Fiers, {M. A.} and Punt, {P. J.} and R. Contreras and M. Penttil{\"a} and {Van Den Hondel}, {C. A.M.J.J.}",
year = "1997",
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Isolation and analysis of functional homologues of the secretion-related SAR1 gene of Saccharomyces cerevisiae from Aspergillus niger and Trichoderma reesei. / Veldhuisen, G.; Saloheimo, M.; Fiers, M. A.; Punt, P. J.; Contreras, R.; Penttilä, M.; Van Den Hondel, C. A.M.J.J.

In: Molecular and General Genetics, Vol. 256, No. 4, 27.11.1997, p. 446-455.

Research output: Contribution to journalArticleScientificpeer-review

TY - JOUR

T1 - Isolation and analysis of functional homologues of the secretion-related SAR1 gene of Saccharomyces cerevisiae from Aspergillus niger and Trichoderma reesei

AU - Veldhuisen, G.

AU - Saloheimo, M.

AU - Fiers, M. A.

AU - Punt, P. J.

AU - Contreras, R.

AU - Penttilä, M.

AU - Van Den Hondel, C. A.M.J.J.

PY - 1997/11/27

Y1 - 1997/11/27

N2 - The Aspergillus niger and Trichoderma reesei genes encoding the functional homologues of the small GTP-binding protein SAR1p, which is involved in the secretion pathway in Saccharomyces cerevisiae, have been cloned and characterised. The A. niger gene (sarA) contains five introns, whereas the T. reesei gene (sar1) has only four. In both cases the first intron is at the same position as the single S. cerevisiae SAR1 intron. The encoded proteins show 70-80% identity to the SAR1 protein. Complementation of S. cerevisiae sar1 and sec12 mutants by expression vectors carrying the A. niger sarA and T. reesei sar1 cDNA clones confirmed that the cloned genes are functional homologues of the S. cerevisiae SAR1 gene. Three mutant alleles of the A. niger sarA gene (D29G, E109K, D29G/E109K), generated by site-directed mutagenesis, revealed a thermosensitive dominant-negative phenotype in the presence of the wild-type sarA allele. This result contrasts with the situation in S. cerevisiae, where similar mutations have a thermosensitive phenotype. Taken together, our results indicate that the sarA gene is involved in an essential function in A. niger.

AB - The Aspergillus niger and Trichoderma reesei genes encoding the functional homologues of the small GTP-binding protein SAR1p, which is involved in the secretion pathway in Saccharomyces cerevisiae, have been cloned and characterised. The A. niger gene (sarA) contains five introns, whereas the T. reesei gene (sar1) has only four. In both cases the first intron is at the same position as the single S. cerevisiae SAR1 intron. The encoded proteins show 70-80% identity to the SAR1 protein. Complementation of S. cerevisiae sar1 and sec12 mutants by expression vectors carrying the A. niger sarA and T. reesei sar1 cDNA clones confirmed that the cloned genes are functional homologues of the S. cerevisiae SAR1 gene. Three mutant alleles of the A. niger sarA gene (D29G, E109K, D29G/E109K), generated by site-directed mutagenesis, revealed a thermosensitive dominant-negative phenotype in the presence of the wild-type sarA allele. This result contrasts with the situation in S. cerevisiae, where similar mutations have a thermosensitive phenotype. Taken together, our results indicate that the sarA gene is involved in an essential function in A. niger.

KW - Complementation

KW - Filamentous fungi

KW - Gene cloning

KW - Protein secretion

KW - Small GTP binding protein

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