Expression of Poplar Chitinase in Tomato Leads to Inhibition of Development in Colorado Potato Beetle

The previously described poplar chitinase, WIN6, is induced during infestation by gypsy moth (Lymantria dispar L.) larvae, thus suggesting a role in defense against insect pests. To test this hypothesis, we produced tomato seedlings infected with a recombinant potato virus X (PVX), which produces WIN6, and tested its insecticidal properties on Colorado potato beetle [CPB; Leptinotarsa decemlineata (Say)], which is a serious pest of tomatoes and other crops. The advantage of PVX is that plant material is ready for insect bioassay within 3–4 weeks of constructing the recombinant virus. Considering that production of transgenic tomato seedlings using Agrobacterium takes at least 6 months, this hastens the rate at which genes can be examined. Upon insect bioassay, only 47% CPB neonates feeding on leaves containing >0.3% w/w WIN6 developed to 2nd instar while 93% of controls reached 2nd instar. To our knowledge this is the first plant chitinase that retards development of an insect pest.

[1]  Jiehua Wang,et al.  Polyphenol oxidase overexpression in transgenic Populus enhances resistance to herbivory by forest tent caterpillar (Malacosoma disstria) , 2004, Planta.

[2]  B. Henrissat,et al.  Classification of chitinases modules. , 1999, EXS.

[3]  T. Congiu,et al.  AcMNPV ChiA protein disrupts the peritrophic membrane and alters midgut physiology of Bombyx mori larvae. , 2004, Insect biochemistry and molecular biology.

[4]  M. Gordon,et al.  Systemic accumulation of specific mRNAs in response to wounding in poplar trees. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[5]  D. Baulcombe,et al.  Potato virus X as a vector for gene expression in plants. , 1992, The Plant journal : for cell and molecular biology.

[6]  P. Faulkner,et al.  Monoclonal antibodies to baculovirus structural proteins: determination of specificities by Western blot analysis. , 1983, Virology.

[7]  G. Blissard,et al.  Identification of a membrane fusion domain and an oligomerization domain in the baculovirus GP64 envelope fusion protein , 1995, Journal of virology.

[8]  C. P. Constabel,et al.  Polyphenol oxidase from hybrid poplar. Cloning and expression in response to wounding and herbivory. , 2000, Plant physiology.

[9]  F. White,et al.  Insect resistance of transgenic tobacco expressing an insect chitinase gene , 1998, Transgenic Research.

[10]  K. Kramer,et al.  Insect chitinases: molecular biology and potential use as biopesticides. , 1997, Insect biochemistry and molecular biology.

[11]  N. Novak,et al.  Epitope tagging: a monoclonal antibody specific for recombinant fusion proteins in plants. , 2003, BioTechniques.

[12]  I. Major,et al.  A Kunitz trypsin inhibitor gene family from trembling aspen (Populus tremuloides Michx.): cloning, functional expression, and induction by wounding and herbivory , 2001, Plant Molecular Biology.

[13]  S. Lawrence,et al.  Chitinase accumulates systemically in wounded poplar trees , 1998 .

[14]  Y. Rahbé,et al.  Transgenic potato plants with enhanced resistance to the peach‐potato aphid Myzus persicae , 1996 .

[15]  D. O'reilly,et al.  Baculovirus expression vectors: a laboratory manual. , 1992 .

[16]  R. Gilbert,et al.  Transgenic potato plants with enhanced resistance to the tomato moth, Lacanobia oleracea: growth room trials , 1997, Molecular Breeding.

[17]  A. Forgash.  Insecticide resistance in the Colorado potato beetle. , 1985 .

[18]  J. Cairney,et al.  A simple and efficient method for isolating RNA from pine trees , 1993, Plant Molecular Biology Reporter.

[19]  I. Chet,et al.  Heterologous chitinase gene expression to improve plant defense against phytopathogenic fungi , 1997, Journal of Industrial Microbiology and Biotechnology.

[20]  N. Novak,et al.  A rapid method for the production and characterization of recombinant insecticidal proteins in plants , 2001, Molecular Breeding.

[21]  S. Lawrence,et al.  A vegetative storage protein homolog is expressed in the growing shoot apex of hybrid poplar , 1997, Planta.

[22]  D. Baulcombe,et al.  Jellyfish green fluorescent protein as a reporter for virus infections. , 1995, The Plant journal : for cell and molecular biology.

[23]  C. Lamb,et al.  Chitinase cDNA cloning and mRNA induction by fungal elicitor, wounding, and infection. , 1988, Plant Physiology.