Growth,development, feeding preference,and food consumption and utilization by tobacco hornworm on tobacco mosaic virus-infected and non-infected tobacco leaves

Summary Plant disease may cause a variety of changes in plants which influence their suitability to insect herbivores. The reported physiological changes in tobacco due to infection by tobacco mosaic virus have the potential to influence the feeding, growth and behavior of the tobacco hornworm,Manduca sexta. However, feeding on foliage of systematically infected plants had no statistically significant effects on development time, mean pupal weight or the amount of foliage consumed by 4th instars. Although an initial preference was exhibited by larvae for healthy leaves after about 72 h no differences were observed. These results do not support the results of previous research on the influence of tobacco mosaic virus-infected plants on the tobacco hornworm.Scientific contribution No. 8130, article No. A-5071, of the Maryland Agricultural Experiment Station, Department of Entomology     

Cyanidin-3-β-glucoside,a newly recognized basis for resistance in cotton to the tobacco budwormheliothis virescens (Fab.) (lepidoptera: Noctuidae)

Summary Cyanidin-3--glucoside was shown to be an important factor of resistance in cottonGossypium hirsutum L. leaves to the tobacco budwormHeliothis virescens (Fab.). This provides a potential basis for achieving insect resistance in non-glanded cotton and other crops infested byHeliothis.In cooperation with the Mississippi Agriculture and Forestry Experiment Station, Mississippi State. Mention of a trademark, proprietary product or vendor does not constitute a guarantee or warranty of the product by the U.S. Department of Agriculture and does not imply its approval to the exclusion of other products or vendors that may also be suitable.     

Signaling versus punching hole: How do Bacillus thuringiensis toxins kill insect midgut cells?

Cry proteins, produced by Bacillus thuringiensis (Bt), are widely used for the control of insect pests in agriculture as spray products or expressed in transgenic crops, such as maize and cotton. Little was known regarding the mechanism of action of these toxins when the first commercial Bt product was introduced fifty years ago. However, research on the mechanism of action over the last two decades has enhanced our knowledge of toxin interaction with membrane receptors and their effects in insect midgut cells. All this information allowed for the rational design of improved toxins with higher toxicity or toxins that overcome insect resistance, which could compromise Bt use and effectiveness in the field. In this review we discuss and evaluate the different models of the mode of action of Cry toxins, including a discussion about the role of various receptors in toxin action. Received 13 June 2008; received after revision 05 November 2008; accepted 11 November 2008     

Hydrogen peroxide protects tobacco from oxidative stress by inducing a set of antioxidant enzymes

Tolerance against oxidative stress generated by high light intensities or the catalase inhibitor aminotriazole (AT) was induced in intact tobacco plants by spraying them with hydrogen peroxide (H₂O₂). Stress tolerance was concomitant with an enhanced antioxidant status as reflected by higher activity and/or protein levels of catalase, ascorbate peroxidase, guaiacol peroxidases, and glutathione peroxidase, as well as an increased glutathione pool. The induced stress tolerance was dependent on the dose of H₂O₂ applied. Moderate doses of H₂O₂ enhanced the antioxidant status and induced stress tolerance, while higher concentrations caused oxidative stress and symptoms resembling a hypersensitive response. In stress-tolerant plants, induction of catalase was 1.5-fold, that of ascorbate peroxidase and glutathione peroxidase was 2-fold, and that of guaiacol peroxidases was approximately 3-fold. Stress resistance was monitored by measuring levels of malondialdehyde, an indicator of lipid peroxidation. The levels of malondialdehyde in all H₂O₂-treated plants exposed to subsequent high light or AT stress were similar to those of unstressed plants, whereas lipid peroxidation in H₂O₂-untreated plants stressed with either high light or AT was 1.5- or 2-fold higher, respectively. Although all stress factors caused increases in the levels of reduced glutathione, its levels were much higher in all H₂O₂-pretreated plants. Moreover, significant accumulation of oxidized glutathione was observed only in plants that were not pretreated with H₂O₂. Extending the AT stress period from 1 to 7 days resulted in death of tobacco plants that were not pretreated with H₂O₂, while all H₂O₂-pretreated plants remained little affected by the prolonged treatment. Thus, activation of the plant antioxidant system by H₂O₂ plays an important role in the induced tolerance against oxidative stress. Received 11 December 2001; received after revision 25 January 2002; accepted 4 February 2002     

Bromocriptine/SKF38393 ameliorates islet dysfunction in the diabetic (db/db) mouse

Dysfunction of pancreatic islets plays a crucial role in the etiology of type II diabetes. Chronic hyperglycaemia or hyperlipidaemia may impair islet function. Previous studies by our laboratory have demonstrated that dopaminergic agonists ameliorated hyperglycaemia and hyperlipidaemia in obese and diabetic rodents. In the present study, we investigated the effect of a treatment with the dopamine D₂ /D₁ receptor agonists (bromocriptine/SKF38393, BC/SKF) on islet dysfunction in db/db mice. Our results show that a 2-week BC/SKF treatment markedly reduced hyperglycaemia and hyperlipidaemia, and significantly improved islet dysfunction demonstrated by an increase of secretagogue-stimulated insulin release from islets of db/db mice to levels observed in islets from lean mice. There was also a fourfold increase of insulin content in the pancreas of BC/SKF-treated db/db mice compared with that in untreated controls. The effect of BC/SKF on islet function cannot be mimicked in pair-fed animals. BC/SKF had no direct stimulatory effect on islet insulin secretion, suggesting BC/SKF treatment improved islet function via an indirect mechanism. This treatment markedly improved the abnormally elevated daily levels of corticosterone, blood glucose and plasma lipids, supporting the view that BC/SKF may affect the neuroendocrine system that in turn regulates peripheral metabolism and thereby improves islet function. Received 3 April 1998; accepted 27 April 1998     

Dictyostelium discoideum cells shed vesicles with associated DNA and vital stain Hoechst 33342

Dictyostelium discoideum cells are highly resis tant to xenobiotics. We previously observed that these primitive eukaryotic cells contain a 170-kDa P-glycoprotein, mediating multidrug resistance in mammalian cells, but nonfunctional in Dictyostelium cells. We show here that D. discoideum cells vitally stained with the DNA-specific dye, Hoechst 33342, release fluorescent material in their culture medium. Electron microscopy and lipid analysis demonstrate the vesicular nature of this material. Moreover, nucleic acids associate with these extracellular vesicles independently of Hoechst vital staining. The main vesicular DNA component exhibits a size >21 kb. Shedding of microvesicles during cell growth is not concomitant with programmed cell death. We propose that these extracellular vesicles are involved in a new cellular resistance mechanism against xenobiotics. Furthermore, since the association of DNA with vesicles occurs in physiological growth conditions and independently of vital staining, the new shedding process might be involved in a more general intercellular mechanism. Received 14 November 1997; received after revision 16 March 1998; accepted 16 March 1998     

Production of functional rat liver PSP protein in Escherichia coli

An efficient Escherichia coli expression system for the production of a perchloric acid-soluble protein (PSP) has been constructed. Complementary DNA encoding PSP was inserted into an inducible bacterial expression vector pGEX-4T-1. After the plasmid introduced into E. coli was expressed by isopropyl 1-thio-β-D-galactopyranoside (IPTG), the recombinant product was purified by glutathione-Sepharose 4B affinity chromatography. The purified product showed the expected NH₂-terminal sequence, but the translation inhibitory activity of this product was 10 times lower compared with that of authentic PSP isolated from rat liver. Received 8 October 1998; received after revision 6 November 1998; accepted 6 November 1998     

Some cellular and molecular properties of abscisic acid: its particular involvement in growing plant roots

Several characteristics of the plant hormone abscisic acid (ABA) are critically discussed, more or less directly, in relation to the extension of root cells. A few topics have been selected some biochemical characteristics of ABA (chemical structure, metabolism), inhibiting-β complex, inhibiting regulators from root caps, endogenous ABA in growing roots (ABA gradients, microsurgical experiments, light effects), applied ABA on elongating roots, ABA and indol-3yl acetic acid (IAA) interactions (root growth, proton extrusion, hormone transport, auxin herbicides), ABA effect on the root cell cycle, ABA and drought cells of elongating roots [water deficit conditions, IAA and jasmonic acid (JA) as ‘stress hormones’ other than ABA, gene expression]. Received 28 January 1998; received after revision 20 April 1998; accepted 21 April 1998     

G proteins as drug targets

The structure and function of heterotrimeric G protein subunits is known in considerable detail. Upon stimulation of a heptahelical receptor by the appropriate agonists, the cognate G proteins undergo a cycle of activation and deactivation; the α-subunits and the βγ-dimers interact sequentially with several reaction partners (receptor, guanine nucleotides and effectors as well as regulatory proteins) by exposing appropriate binding sites. For most of these domains, low molecular weight ligands have been identified that either activate or inhibit signal transduction. These ligands include short peptides derived from receptors, G protein subunits and effectors, mastoparan and related insect venoms, modified guanine nucleotides, suramin analogues and amphiphilic cations. Because compounds that act on G proteins may be endowed with new forms of selectivity, we propose that G protein subunits may therefore be considered as potential drug targets. Received 18 September 1998; received after revision 6 November 1998; accepted 11 November 1998     

Lack of conversion of C29-phytosterols to cholesterol in the khapra beetle,Trogoderma granarium Everts

Summary Metabolic studies in which³H-sitosterol,³H-stigmasterol, and¹⁴C-desmosterol were administered by feeding and injection to the khapra beetle,Trogoderma granarium Everts, provided strong evidence that this insect is unable to dealkylate phytosterols and convert them to cholesterol.     

Identifying the conformational state of bi-liganded haemoglobin

While most researchers agree on the global features of cooperative ligand binding to haemoglobin (Hb), the internal mechanisms remain open to debate. This is not due to inaccurate measurements, but is rather a consequence of the cooperative ligand binding that decreases the equilibrium populations of the partially liganded states and makes observation of the transitions between these substates more difficult. For example, the equilibrium population of the doubly liganded tetramers is typically less than 5% of the total Hb. As a result many models with widely varying mechanisms may fit the oxygen equilibrium curve, but may not be consistent with observations of other parameters, such as ligand-binding kinetics or subunit association equilibria. The wide range of methods and models has led to divergent conclusions about the properties of specific substates. One notable debate concerns the properties of the doubly liganded forms. The simple two-state model predicts a shift in the allosteric equilibrium based on the number of ligands bound, but not on their distribution within the tetramer. From studies of dimer-tetramer equilibria of various pure and hybrid forms, it was concluded that a tetramer with two ligands bound on the same α β dimer (species 21, an asymmetric hybrid) shows an enhanced tetramer stability, similar to singly liganded Hb, relative to the other three types of doubly liganded tetramers which resemble the triply liganded forms [Ackers et al. (1992), Science 255: 54–63]. The implications of this model and the relevant experiments will be reviewed here. Received 27 April 1998; received after revision 17 July 1998; accepted 10 August 1998     

Effects of feeding on healthy and diseased corn plants on a vector and on a non-vector insect

Insect-transmitted plant pathogens can have positive, negative or no effects on the vector insect. Effects could be direct (caused by the pathogen) or indirect (mediated by changes in the plant). Pathogen effects on non-vector insects are less well known.Peregrinus maidis (Ashmead) (Homoptera: Delphacidae), an insect that is not a vector of the corn stunt spiroplasma (CSS), weighed less 15 days after feeding on CSS-infected corn plants relative to insects feeding on healthy corn plants. Fecundity of non-vector insects that were removed from diseased plants was higher than for insects coming from healthy plants. ForDalbulus maidis (DeLong & Wolcott) (Homoptera: Cicadellidae), an insect that transmits CSS, there were no differences in weight, longevity, eggs per day, weekly or total fecundity after feeding on healthy or on CSS-infected corn plants. Significant differences in some phloem amino acids were detected between healthy and CSS-infected plants. Infected plants also showed an increased phloem acidity as disease symptoms progressed. Feeding on CSS-infected corn plants by an insect that does not vector the pathogen infecting the plant can have favorable consequences as evidenced by an increased fecundity.Scientific aritcle no. 8819, contribution no. A-6605 of the Maryland Agricultural Experiment Station.     

Selective herbivory on mosaic leaves of variegatedAcer pseudoplatanus

Summary The color mosaicism was observed to correlate with selective herbivory on different leaf areas of individual leaves of the variegated sycamoreAcer pseudoplatanus. The mosaicism affected the content of nutrients and of phenolic compounds and also the mechanical properties of the different leaf areas; this offers an explanation for the observed selective feeding by insect herbivores.Acknowledgments. We wish to thank S. Neuvonen for critical comments and C. Grapes for checking the language. The work was supported by the Academy of Finland and J. Salo Foundation.     

Synthesis and metabolism of vertebrate-type steroids by tissues of insects: A critical evaluation

Summary This review covers the synthesis and the metabolism of vertebrate-type steroids (progesterone, testosterone, estradiol, corticosteroids) by insect tissues and discusses the significance of the reactions for insect physiology. Biosynthesis of vertebrate-type steroids from cholesterol hitherto has been demonstrated in only two insect species, i.e. the water beetleAcilius sulcatus (Coleoptera) and the tobacco hornwormManduca sexta (Lepidoptera). InAcilius, steroid synthesis is associated with exosecretion (chemical defense). Nothing, however, is known about a physiological role of the C₂₁ steroid conjugate present in ovaries and eggs ofManduca. No synthesis of vertebrate-type steroids was observed in any other insect investigated to date. Most metabolic conversions of steroids by insects concerned oxidoreduction of oxygen groups (hydroxysteroid dehydrogenase activity) and (polar and apolar) conjugate formation. All important enzymatic steps involved in synthesis and catabolism, as known from studies with tissues of vertebrates, were not, or hardly observed. The conclusion is drawn that typical vertebrate-type (C₂₁, C₁₉ and C₁₈) steroids probably do not act as physiologically active substances in insects.     

Pycnogenol enhances immune and haemopoietic functions in senescence-accelerated mice

Pycnogenol (procyanidin extracted from Pinus maritima) has been shown to be a potent free radical scavenger and an antioxidant phytochemical. The effects of pycnogenol on immune and haemopoietic dysfunction in senescence-accelerated mice (SAM), as a murine model of accelerated ageing, were determined. SAMP8, a strain of senile-prone mice, exhibit learning and memory deficits, immunodeficiency and dysfunction of the haemopoietic system. Oral feeding with pycnogenol for 2 months significantly improved their T- and B-cell function. Pycnogenol also augmented the proliferative capacity of haemopoietic progenitors of bone marrow in SAMP8. These data suggest that pycnogenol may be useful for either retardation or restoration of parameters associated with ageing. Received 3 July 1998; accepted 28 July 1998     

Effects of the adenylate cyclase activator forskolin and its inactive derivative 1,9-dideoxyforskolin on insect cytochrome P-450 dependent steroid hydroxylase activity

The adenylate cyclase activator forskolin and its pharmacologically inactive derivative 1,9-dideoxyforskolin were found to inhibit in a dose-dependent fashion the ecdysone 20-monooxygenase activity associated with wandering stage larvae ofDrosophila melanogaster and fat body and midgut from last instar larvae of the tobacco hornworm,Manduca sexta. The concentrations of these labdane diterpenes required to elicit a 50% inhibition of the cytochrome P-450 dependent steroid hydroxylase activity in the insect tissues ranged from approximately 5×10^–6 to 5×10^–4 M.     

Growth inhibitory,insecticidal and antifeedant effects of some antileukemic and cytotoxic quassinoids on two species of agricultural pests

Summary Several quassinoids, obtained by isolation and derivatization fromSimaba multiflora andSoulamea soulameoides, were evaluated for growth inhibitory and insecticidal effects against the tobacco budworm (Heliothis virescens) and for antifeedant effects againstH. virescens and the fall armyworm (Spodoptera frugiperda). The relative activity of the quassinoids as insect growth inhibitors generally paralleled their known relative potency as antileukemic and cytotoxic agents.Phytochemical aspects of this work were supported by Contract CM-97295 with the Division of Cancer Treatment, National Cancer Institute, National Institutes of Health, Bethesda, Md. Insects were kindly supplied by the agency of the United States Department of Agriculture at Brownsville, TX.     

In pre-sterol carrier protein 2 (SCP2) in solution the leader peptide 1 – 20 is flexibly disordered, and residues 21 – 143 adopt the same globular fold as in mature SCP2

The preform of the rabbit sterol carrier protein 2 (pre-rSCP2) was cloned, the uniformly ¹⁵N-labelled protein expressed in Escherichia coli and studied by three-dimensional ¹⁵N-resolved nuclear magnetic resonance spectroscopy. In spite of its low solubility in aqueous solution of only ∼0.3 mM, sequential ¹⁵N and ¹H backbone resonance assignments were obtained for 105 out of the 143 residues. From comparison of the sequential and medium-range nuclear Overhauser effects (NOEs) in the two proteins, all regular secondary structures previously determined in mature human SCP2 (hSCP2) [Szyperski et al. (1993) FEBS Lett. 335: 18–26] were also identified in pre-rSCP2. Near-identity of the backbone ¹⁵N and ¹H chemical shifts and 1 : 1 correspondence of 24 long-range NOEs to backbone amide groups in the two proteins show that the residues 21 – 143 adopt the same globular fold in pre-rSCP2 and mature hSCP2. The N-terminal 20-residue leader peptide of pre-rSCP2 is flexibly disordered in solution and does not observably affect the conformation of the polypeptide segment 21 – 143. Received 11 May 1998; accepted 15 May 1998     

MDR1, cholesterol certification and cell growth: a comparative study in normal and multidrug-resistant KB cell lines

The product of the MDR1 gene (P-gp) has been implicated in the transport of cholesterol from plasma membrane to endoplasmic reticulum for esterification. In previous studies on leukemia cell lines, we suggested that cholesterol esterification may regulate the rate of cell growth and that the MDR1 gene might be involved in this process by modulating intracellular cholesterol esters levels. To further investigate this matter, the rate of cell growth, cholesterol metabolism, expression of the MDR1 gene, and P-gp activity were compared in KB cell lines displaying differences in expression and function of P-gp (drug-sensitive phenotype versus MDR phenotype). The rate of cell growth correlated with cholesterol esterification in all KB cell lines, whereas the over-expression of MDR1 observed in the MDR cell lines was not always associated with an increased capacity of cells to esterify cholesterol. Two known inhibitors of P-gp activity, progesterone and verapamil, strongly inhibited both cholesterol esterification and cell proliferation in all KB cell lines, but they affected intracellular accumulation of labeled vinblastine only in MDR cell lines. These results further support a role for cholesterol esters in the regulation of cell growth and suggest that the P-gp expressed in MDR KB cells is not involved in the general process leading to cholesterol esterification. Received 14 February 2000; received after revision 10 April 2000; accepted 8 May 2000