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 language | English |
|---|---|
| Pages (from-to) | 446-455 |
| Number of pages | 10 |
| Journal | Molecular and General Genetics |
| Volume | 256 |
| Issue number | 4 |
| DOIs | |
| Publication status | Published - 27 Nov 1997 |
| MoE publication type | A1 Journal article-refereed |
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Keywords
- Complementation
- Filamentous fungi
- Gene cloning
- Protein secretion
- Small GTP binding protein
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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 journal › Article › Scientific › peer-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
UR - http://www.scopus.com/inward/record.url?scp=0030685697&partnerID=8YFLogxK
U2 - 10.1007/s004380050588
DO - 10.1007/s004380050588
M3 - Article
C2 - 9393442
AN - SCOPUS:0030685697
VL - 256
SP - 446
EP - 455
JO - Molecular Genetics and Genomics
JF - Molecular Genetics and Genomics
SN - 1617-4615
IS - 4
ER -