MEK/ERK MAP kinase limits poly I:C-induced antiviral gene expression in RAW264.7 macrophages by reducing interferon-beta expression.

Toll-like receptor 3 (TLR3) recognizes viral double-stranded RNA (or the synthetic dsRNA analog poly I:C) and induces a signal transduction pathway that results in activation of transcription factors that induce expression of antiviral genes including type I interferon (IFN-I). Secreted IFN-I positively feeds back to amplify antiviral gene expression. In this report, we study the role of MEK/ERK MAP kinase in modulating antiviral gene expression downstream of TLR3. We find MEK/ERK is a negative regulator of antiviral gene expression by limiting expression of IFN-ß. However, MEK/ERK does not limit antiviral responses downstream of the type I interferon receptor. These findings provide insights into regulatory mechanisms of antiviral gene expression and reveal potential targets for modulating antiviral immunity.

Organophosphorus hydrolase: a multifaceted plant genetic marker which is selectable, scorable, and quantifiable in whole seed.

Organophosphorus hydrolase (OPH, EC 3.1.8.1) provides a novel function as an alternative genetic marker system for use in many types of plant transformations. OPH is a high-capacity hydrolase with multiple organophosphorus substrates, many of which are neurotoxins and thus used extensively as pesticides. This spectrum of organophosphates includes compounds that are phytotoxic as well as those that are hydrolyzed to products that are easily detected visually without significant disruption of plant health. This dichotomy gives OPH the features of both a selectable marker as well as that of a scorable marker system, and these characteristics have been tested at several stages during the plant transformation and regeneration process. Finally, it is possible to quantify hydrolytic activity in the seed without interfering with its subsequent growth and regeneration.

Identification and properties of insect resistance-associated maize anionic peroxidases.

Previous studies with transgenic plants have indicated a tobacco anionic peroxidase can confer enhanced resistance to a variety of insects when expressed in different plant species. Tissue that expresses high levels of this enzyme often browns rapidly when damaged. Maize roots damaged under sterile conditions browned and had an anionic peroxidase induced. When introduced biolistically, maize callus transformants expressing a maize peroxidase gene with a predicted isoelectric point of ca. 5.1 produced browner callus compared to a corresponding beta-glucuronidase (GUS) transformant as callus aged. Higher production of only one isozyme of ca. pI 4.5 was noted. When the callus was fed to two maize pest caterpillar species, growth rates were slower (as reflected by weights) relative to the GUS callus. Based on examination of published information and electrophoretic properties, this gene appears to code for Px11, a peroxidase isozyme that is primarily produced in root tissue and callus. When sequence of the gene in several inbreds was examined, coding variations were noted, and abilities to utilize ferulic and p-coumaric acids differed. These coding differences may influence the ability of corresponding forms of the peroxidase to promote resistance. In addition to potential use in marker assisted breeding, enhanced expression of this anionic peroxidase through breeding or genetic engineering may lead to enhanced insect or disease resistance.

Oral toxicity of beta-N-acetyl hexosaminidase to insects.

Insect chitin is a potential target for resistance plant proteins, but plant-derived chitin-degrading enzymes active against insects are virtually unknown. Commercial beta-N-acetylhexosaminidase (NAHA), a chitin-degrading enzyme from jack bean Canavalia ensiformis, caused significant mortality of fall armyworm Spodoptera frugiperda larvae at 75 microg/gm, but no significant mortality was noted with Aspergillus niger NAHA. Maize Zea mays callus transformed to express an Arabidopsis thaliana clone that putatively codes for NAHA caused significantly higher mortality of cigarette beetle Lasioderma serricorne larvae and significantly reduced growth rates (as reflected by survivor weights) of S. frugiperda as compared to callus that expressed control cDNAs. Tassels from model line Hi-II maize (Z. mays) plants transformed with the NAHA gene fed to S. frugiperda caused significantly higher mortality than tassels transformed to express glucuronidase; mortality was significantly correlated with NAHA expression levels detected histochemically. Leaf disks from inbred Oh43 maize plants transformed with the NAHA gene on average had significantly less feeding by caterpillars than null transformants. Leaf disks of Oh43 transformants caused significant mortality of both S. frugiperda and corn earworm Helicoverpa zea larvae, which was associated with higher expression levels of NAHA detected by isoelectric focusing, histochemically, or with antibody. Overall, these results suggest that plant NAHA has a role in insect resistance. Introduction of NAHA genes or enhancement of activity through breeding or genetic engineering has the potential to significantly reduce insect damage and thereby indirectly reduce mycotoxins that are harmful to animals and people.

Relative activity of a tobacco hybrid expressing high levels of a tobacco anionic peroxidase and maize ribosome-inactivating protein against Helicoverpa zea and Lasioderma serricorne.

Tobacco (Nicotiana tabacum) plants grown from seed obtained by crossing a tobacco line that expressed an activated maize ribosome-inactivating protein (RIP) with a line that overexpressed tobacco anionic peroxidase were tested for their effects on corn earworm Helicoverpa zea and cigarette beetle Lasioderma serricorne larvae as compared to the wild-type plant cross. Significant feeding reductions were noted for transgenic plants expressing both resistance proteins as compared to wild-type plants for both H. zea and L. serricorne. Significant increases in mortality were also noted for those insects fed on the transgenic cross as compared to wild-type plants in some cases. Levels of both peroxidase and maize RIP were significantly higher in transgenic as compared to wild-type plants (which did not produce maize RIP). The degree of feeding was significantly negatively correlated with the level of RIP or peroxidase individually.