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Applied and Environmental Microbiology, February 2002, p. 532-538, Vol. 68, No. 2
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.2.532-538.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Osmoregulation and Fungicide Resistance: the Neurospora crassa os-2 Gene Encodes a HOG1 Mitogen-Activated Protein Kinase Homologue{dagger}

Yan Zhang,1 Randy Lamm,1 Christian Pillonel,1 Stephen Lam,1 and Jin-Rong Xu2*

Syngenta Agribusiness Biotechnology Research Inc., Research Triangle Park, North Carolina 27709,1 Department of Botany and Plant Pathology, Purdue University, West Lafayette, Indiana 479072

Received 21 August 2001/ Accepted 14 November 2001

Neurospora crassa osmosensitive (os) mutants are sensitive to high osmolarity and therefore are unable to grow on medium containing 4% NaCl. We found that os-2 and os-5 mutants were resistant to the phenylpyrrole fungicides fludioxonil and fenpiclonil. To understand the relationship between osmoregulation and fungicide resistance, we cloned the os-2 gene by using sib selection. os-2 encodes a putative mitogen-activated protein (MAP) kinase homologous to HOG1 and can complement the osmosensitive phenotype of a Saccharomyces cerevisiae hog1 mutant. We sequenced three os-2 alleles and found that all of them were null with either frameshift or nonsense point mutations. An os-2 gene replacement mutant also was generated and was sensitive to high osmolarity and resistant to phenylpyrrole fungicides. Conversely, os-2 mutants transformed with the wild-type os-2 gene could grow on media containing 4% NaCl and were sensitive to phenylpyrrole fungicides. Fludioxonil stimulated intracellular glycerol accumulation in wild-type strains but not in os-2 mutants. Fludioxonil also caused wild-type conidia and hyphal cells to swell and burst. These results suggest that the hyperosmotic stress response pathway of N. crassa is the target of phenylpyrrole fungicides and that fungicidal effects may result from a hyperactive os-2 MAP kinase pathway.

* Corresponding author. Mailing address: Department of Botany and Plant Pathology, Purdue University, West Lafayette, IN 47907. Phone: (765) 496-6918. Fax: (765) 494-0363. E-mail: xu{at}btny.purdue.edu.

{dagger} Journal article no. 16663 of the Purdue University Agricultural Experiment Station.

Applied and Environmental Microbiology, February 2002, p. 532-538, Vol. 68, No. 2
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.2.532-538.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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