Title
A novel screening method for cell wall mutants in Aspergillus niger identifies UDP-galactopyranose mutase as an important protein in fungal cell wall biosynthesis
Author
Damveld, R.A.
Franken, A.
Arentshorst, M.
Punt, P.J.
Klis, F.M.
van den Hondel, C.A.M.J.J.
Ram, A.F.J.
TNO Kwaliteit van Leven
Publication year
2008
Abstract
To identify cell wall biosynthetic genes in filamentous fungi and thus potential targets for the discovery of new antifungals, we developed a novel screening method for cell wall mutants. It is based on our earlier observation that the Aspergillus niger agsA gene, which encodes a putative a-glucan synthase, is strongly induced in response to cell wall stress. By placing the agsA promoter region in front of a selectable marker, the acetamidase (amdS) gene of A. nidulans, we reasoned that cell wall mutants with a constitutively active cell wall stress response pathway could be identified by selecting mutants for growth on acetamide as the sole nitrogen source. For the genetic screen, a strain was constructed that contained two reporter genes controlled by the same promoter: the metabolic reporter gene PagsA-amdS and PagsA-H2B-GFP, which encodes a GFP-tagged nuclear protein. The primary screen yielded 161 mutants that were subjected to various cell wall-related secondary screens. Four calcofluor white-hypersensitive, osmotic-remediable thermosensitive mutants were selected for complementation analysis. Three mutants were complemented by the same gene, which encoded a protein with high sequence identity with eukaryotic UDP-galactopyranose mutases (UgmA). Our results indicate that galactofuranose formation is important for fungal cell wall biosynthesis and represents an attractive target for the development of antifungals. Copyright © 2008 by the Genetics Society of America.
Subject
Biology
Biotechnology
Acetamide
Alpha glucan synthase
Green fluorescent protein
Histone H2B
Mutase
Nuclear protein
Uridine diphosphate
Uridine diphosphate galactopyranose mutase
Article
Aspergillus niger
Biosynthesis
Cell wall
Controlled study
DNA sequence
Gene deletion
Gene expression
Mutagenesis
Nonhuman
PagsA gene
Priority journal
Reporter gene
Screening test
Temperature sensitive mutant
Aspergillus niger
Cell Wall
Fungal Proteins
Gene Deletion
Intramolecular Transferases
Mutagenesis
Mutation
Recombination, Genetic
Aspergillus niger
Eukaryota
Fungi
To reference this document use:
http://resolver.tudelft.nl/uuid:8d77085e-bd76-4bff-9dc1-5471369b2910
DOI
https://doi.org/10.1534/genetics.107.073148
TNO identifier
240644
ISSN
0016-6731
Source
Genetics, 178 (2), 873-881
Document type
article