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Applied and Environmental Microbiology, August 2004, p. 4989-4995, Vol. 70, No. 8
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.8.4989-4995.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

VdNEP, an Elicitor from Verticillium dahliae, Induces Cotton Plant Wilting

Jian-Ying Wang,1 Yu Cai,1 Jin-Ying Gou,1 Ying-Bo Mao,1 Yan-Hua Xu,1 Wei-Hong Jiang,2 and Xiao-Ya Chen1*

National Key Laboratory of Plant Molecular Genetics,1 Laboratory of Molecular Microbiology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, People's Republic of China2

Received 15 January 2004/ Accepted 26 April 2004

Verticillium wilt is a vascular disease of cotton. The causal fungus, Verticillium dahliae, secretes elicitors in culture. We have generated ~1,000 5'-terminal expressed sequence tags (ESTs) from a cultured mycelium of V. dahliae. A number of ESTs were found to encode proteins harboring putative signal peptides for secretion, and their cDNAs were isolated. Heterologous expression led to the identification of a protein with elicitor activities. This protein, named V. dahliae necrosis- and ethylene-inducing protein (VdNEP), is composed of 233 amino acids and has high sequence identities with fungal necrosis- and ethylene-inducing proteins. Infiltration of the bacterially expressed His-VdNEP into Nicotiana benthamiana leaves resulted in necrotic lesion formation. In Arabidopsis thaliana, the fusion protein also triggered production of reactive oxygen species and induced the expression of PR genes. When added into suspension cultured cells of cotton (Gossypium arboreum), the fusion protein elicited the biosynthesis of gossypol and related sesquiterpene phytoalexins at low concentrations, and it induced cell death at higher concentrations. On cotton cotyledons and leaves, His-VdNEP induced dehydration and wilting, similar to symptoms caused by a crude preparation of V. dahliae elicitors. Northern blotting showed a low level of VdNEP expression in the mycelium during culture. These data suggest that VdNEP is a wilt-inducing factor and that it participates in cotton-V. dahliae interactions.

* Corresponding author. Mailing address: National Key Laboratory of Plant Molecular Genetics, Institute of Plant Physiology and Ecology, Chinese Academy of Sciences, 300 Fenglin Rd., Shanghai 200032, People's Republic of China. Phone: 86-21-54924033. Fax: 86-21-54924015. E-mail: xychen{at}sibs.ac.cn.

Applied and Environmental Microbiology, August 2004, p. 4989-4995, Vol. 70, No. 8
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.8.4989-4995.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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