A Unique 33-kD Cysteine Proteinase Accumulates in Response to Larval Feeding in Maize Genotypes Resistant to Fall Armyworm and Other Lepidoptera

Abstract

Plants respond to insect feeding with a number of defense mechanisms. Using maize genotypes derived from Antiquan germ plasm that are resistant to Lepidoptera, we have demonstrated that a unique 33-kD cysteine proteinase accumulates in the whorl in response to larval feeding. The abundance of the proteinase increased dramatically at the site of larval feeding after 1 hr of infestation and continued to accumulate for as long as 7 days. The 33-kD cysteine proteinase was most abundant in the yellow-green portion of the whorl-the normal site of larval feeding and the tissue that has the greatest inhibitory effect on larval growth in bioassays. The proteinase was expressed in response to wounding and was found in senescent leaves. It may be a marker of programmed cell death. The gene coding for the proteinase, mir1 , has been transformed into Black Mexican Sweet callus. When larvae were reared on callus expressing the proteinase, their growth was inhibited ‫ف‬ 60 to 80%. The expression of a cysteine proteinase, instead of a cysteine proteinase inhibitor, may be a novel insect defense mechanism in plants. INTRODUCTIONOver the past 25 years, maize inbreds resistant to feeding by larvae of numerous lepidopteran species have been developed from Antiguan germ plasm (Williams and Davis, 1982;Williams et al., 1990a). Inbreds derived from this germ plasm (Mp704 and Mp708) are resistant to feeding by fall armyworm ( Spodoptera frugiperda ), southwestern corn borer ( Diatraea grandiosella), European corn borer ( Ostinia nubilalis ), sugarcane borer ( D. saccharalis), tobacco budworm ( Heliothis virescens ), corn earworm ( Helicoverpa zea ), and other Lepidoptera. Fall armyworm larvae feed extensively on whorl leaf tissue, often resulting in crop losses. Genetic and quantitative trait loci analyses indicate that resistance to these Lepidoptera is a quantitative trait regulated by several genes (Williams et al., 1989;Khairallah et al., 1998). Traits such as high hemicellulose content, low protein content, and leaf toughness appear to be correlated with reduced larval growth (Williams et al., 1998). No studies have indicated conclusively that secondary products contribute to the resistance, but two-dimensional gel electrophoresis has indicated that the presence of 36-and 21-kD proteins in the whorl may be predictive of resistance (Callahan et al., 1992).Bioassays in which fall armyworm larvae are reared on lyophilized whorl tissues indicate that larvae reared on resistant material weigh ‫ف‬ 50% less than those reared on susceptible material (Williams et al., 1990b). Larvae reared on lyophilized whorl tissue from resistant genotypes are smaller, grow more slowly, and pupate later than those reared on similar material from susceptible genotypes (Chang et al., 2000). The major effect of this germplasm is to slow larval growth and development and to increase the amount of time larvae are vulnerable to predators and parasites.The same phenotype, a 50% reduction in larval growth, is apparent when larvae are reared on nonfriable callus ...

Mechanisms of sugarcane response to herbivory

“…Its potential application to plant protection against insects is being investigated, particularly against the sugarcane borer, D. saccharalis. In agreement with Pechan et al (2000), we have found that mir1 is highly expressed in sugarcane callus, although higher expression levels were observed in seeds, root transition zone, stem and plants infected with H. rubrisubalbicans and A. diazotroficans (Figure 2d). …”

“…Another interesting insect-resistance gene from a resistant inbred maize line has recently been isolated and characterized by Pechan et al (2000), the gene product being a 33-kDa cysteine proteinase (encoded by mir1 gene) which inhibits the growth of a wide range of lepidopteran larvae. Pechan et al suggest that this might be a novel insect defense mechanism in plants, although its mode of action is still unclear.…”

Enzymatic and Structural Characterization of the Major Endopeptidase in the Venus Flytrap Digestion Fluid

“…The Z-FR-AMC substrate was used for kinetic profiling either at 0.25 mM or in a dilution series to determine kinetic constants. The catalytic competence of dionain-1 at different temperatures (4,20,30,40,50,60,70, and 80°C) was probed by end point fluorescence after 15-min incubation of dionain-1 in preheated 50 mM sodium acetate, pH 5.5, assay buffer containing 2 mM DTT and 0.25 mM Z-FR-AMC. The reaction was stopped by the addition of 1 mM E-64 and incubation for 10 min at room temperature before measuring total fluorescence.…”

Intraplant communication in maize contributes to defense against insects

“…18 The maize genotype, Mp708, developed by classical plant breeding is remarkably resistant to different feeding styles of herbivore pests. 12,[19][20][21] Previous studies have shown that Mp708 provides resistance to caterpillars by rapidly accumulating Maize insect resistance1-Cysteine Protease (Mir1-CP), a papain-like protease. 19 Furthermore, it has been shown that Mir1-CP is localized in maize vascular tissues.…”

“…12,[19][20][21] Previous studies have shown that Mp708 provides resistance to caterpillars by rapidly accumulating Maize insect resistance1-Cysteine Protease (Mir1-CP), a papain-like protease. 19 Furthermore, it has been shown that Mir1-CP is localized in maize vascular tissues. 22 Interestingly, Mir1-CP, a phloem-mobile protein, accumulates in roots in response to foliar feeding by FAW.…”