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A Method for the Selection of Deletion Mutations in the L-Proline Catabolism Gene Cluster of Aspergillus nidulans

Published online by Cambridge University Press:  14 April 2009

Herbert N. Arst Jr ,
Susan A. Jones  and
Christopher R. Bailey
Herbert N. Arst Jr*
Affiliation:
Department of Genetics, University of Cambridge, Downing Street, Cambridge CB2 3EH, England Department of Genetics, Ridley Building, The University, Newcastle upon Tyne NE1 7RU, England
Susan A. Jones
Affiliation:
Department of Genetics, University of Cambridge, Downing Street, Cambridge CB2 3EH, England
Christopher R. Bailey
Affiliation:
Department of Genetics, University of Cambridge, Downing Street, Cambridge CB2 3EH, England
*
* Address correspondence to this author (at Newcastle address).
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Interest in the selection of mutations affecting L-proline catabolism in Aspergillus nidulans is heightened by the involvement of one of the very few examples of a cluster of functionally related genes in an eukaryote and by an increasing awareness of the biological phenomena in which proline and proline catabolism participate. The sasA-60 (semialdehyde sensitive) mutation in A. nidulans results in toxicity of catabolic precursors of L-glutamic γ-semialdehyde (or its internal Schiff base L-Δ1-pyrroline-5-carboxylate) and succinic semialdehyde, apparently without affecting the catabolic pathways concerned. As sasA-60 is unlinked to the prn gene cluster, specifying the gene products necessary for L-proline catabolism and as L-proline, a precursor of L-glutamic γ-semialdehyde, is highly toxic to sasA-60 strains, this forms the basis of a powerful positive selection technique for obtaining a number of types of prn mutations. Many of these prn mutations can be directly classified according to the gene product(s) affected on the basis of growth phenotype with respect to L-arginine and L-ornithine utilization, proline-dependent resistance to certain toxic amino acid analogues and effect on supplementation of proline auxotrophies. The availability of both a positive selection technique and an extensive nutritional screening system has enabled the identification of fourteen spontaneous deletion mutations, recognized as extending into the prnB gene, specifying the principal L-proline permease, and into at least one other prn gene. These deletion mutations have been partially characterized both genetically and biochemically. In particular their use has greatly facilitated fine-structure mapping of the prn cluster and aided studies of the regulation of prn gene expression.

Type
Research Article
Information
Genetics Research , Volume 38 , Issue 2 , October 1981 , pp. 171 - 195
Copyright
Copyright © Cambridge University Press 1981

References

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