Transgenic cereals: current status and future prospects

This review summarises the history of transgenic (GM) cereals, principally maize, and then focuses on the scientific literature published in the last two years. It describes the production of GM cereals with modified traits, divided into input traits and output traits. The first category includes herbicide tolerance and insect resistance, and resistance to abiotic and biotic stresses; the second includes altered grains for starch, protein or nutrient quality, the use of cereals for the production of high value medical or other products, and the generation of plants with improved efficiency of biofuel production. Using data from field trial and patent databases the review considers the diversity of GM lines being tested for possible future development. It also summarises the dichotomy of response to GM products in various countries, describes the basis for the varied public acceptability of such products, and assesses the development of novel breeding techniques in the light of current GM regulatory procedures.

Claviceps purpurea expressing polygalacturonases escaping PGIP inhibition fully infects PvPGIP2 wheat transgenic plants but its infection is delayed in wheat transgenic plants with increased level of pectin methyl esterification

Claviceps purpurea is a biotrophic fungal pathogen of grasses causing the ergot disease. The infection process of C. purpurea on rye flowers is accompanied by pectin degradation and polygalacturonase (PG) activity represents a pathogenicity factor. Wheat is also infected by C. purpurea and we tested whether the presence of polygalacturonase inhibiting protein (PGIP) can affect pathogen infection and ergot disease development. Wheat transgenic plants expressing the bean PvPGIP2 did not show a clear reduction of disease symptoms when infected with C. purpurea. To ascertain the possible cause underlying this lack of improved resistance of PvPGIP2 plants, we expressed both polygalacturonases present in the C. purpurea genome, cppg1 and cppg2 in Pichia pastoris. In vitro assays using the heterologous expressed PGs and PvPGIP2 showed that neither PG is inhibited by this inhibitor. To further investigate the role of PG in the C. purpurea/wheat system, we demonstrated that the activity of both PGs of C. purpurea is reduced on highly methyl esterified pectin. Finally, we showed that this reduction in PG activity is relevant in planta, by inoculating with C. purpurea transgenic wheat plants overexpressing a pectin methyl esterase inhibitor (PMEI) and showing a high degree of pectin methyl esterification. We observed reduced disease symptoms in the transgenic line compared with null controls. Together, these results highlight the importance of pectin degradation for ergot disease development in wheat and sustain the notion that inhibition of pectin degradation may represent a possible route to control of ergot in cereals.

The trafficking pathway of a wheat storage protein in transgenic rice endosperm

Background and Aims The trafficking of proteins in the endoplasmic reticulum (ER) of plant cells is a topic of considerable interest since this organelle serves as an entry point for proteins destined for other organelles, as well as for the ER itself. In the current work, transgenic rice was used to study the pattern and pathway of deposition of the wheat high molecular weight (HMW) glutenin sub-unit (GS) 1Dx5 within the rice endosperm using specific antibodies to determine whether it is deposited in the same or different protein bodies from the rice storage proteins, and whether it is located in the same or separate phases within these. Methods The protein distribution and the expression pattern of HMW sub-unit 1Dx5 in transgenic rice endosperm at different stages of development were determined using light and electron microscopy after labelling with antibodies. Key results The use of HMW-GS-specific antibodies showed that sub-unit 1Dx5 was expressed mainly in the sub-aleurone cells of the endosperm and that it was deposited in both types of protein body present in the rice endosperm: derived from the ER and containing prolamins, and derived from the vacuole and containing glutelins. In addition, new types of protein bodies were also formed within the endosperm cells. Conclusions The results suggest that the HMW 1Dx5 protein could be trafficked by either the ER or vacuolar pathway, possibly depending on the stage of development, and that its accumulation in the rice endosperm could compromise the structural integrity of protein bodies and their segregation into two distinct populations in the mature endosperm.

The extent to which potential benefits to EU farmers of adopting transgenic crops are reduced by cost of compliance with coexistence regulations

This article forecasts the extent to which the potential benefits of adopting transgenic crops may be reduced by costs of compliance with coexistence regulations applicable in various member states of the EU. A dynamic economic model is described and used to calculate the potential yield and gross margin of a set of crops grown in a selection of typical rotation scenarios. The model simulates varying levels of pest, weed, and drought pressures, with associated management strategies regarding pesticide and herbicide application, and irrigation. We report on the initial use of the model to calculate the net reduction in gross margin attributable to coexistence costs for insect-resistant (IR) and herbicide-tolerant (HT) maize grown continuously or in a rotation, HT soya grown in a rotation, HT oilseed rape grown in a rotation, and HT sugarbeet grown in a rotation. Conclusions are drawn about conditions favoring inclusion of a transgenic crop in a crop rotation, having regard to farmers’ attitude toward risk.

Detection of tobacco mosaic tobamovirus in cigarettes through RT-PCR

TMV was tried to recover from a variety of branded cigarettes and cigars. Tobacco from six different brands of cigarettes and cigar were processed and reverse transcriptase polymerase chain reaction was employed for the detection of TMV. RTPCR confirmed the presence of TMV in tobacco from one brand of cigarette and one brand of cigar. Bean plants (Phaseolus vulgaris) were inoculated manually with tobacco sap of cigarettes resulting in the production of localized disease lesions. Together, these results showed that tobacco used to make cigarettes and cigars can function as an effective disease vector, potentially aiding the movement of infectious TMV between countries. This is an important finding prompting a need to test smoking tobacco for other virus particles that infect tobacco plants and survive processing as well as considering biosecurity measures to limit virus transmission

Fitness of transgenic Anopheles stephensi mosquitoes expressing the SM1 peptide under the control of a vitellogenin promoter

Three transgenic Anopheles stephensi lines were established that strongly inhibit transmission of the mouse malaria parasite Plasmodium berghei. Fitness of the transgenic mosquitoes was assessed based on life table analysis and competition experiments between transgenic and wild-type mosquitoes. Life table analysis indicated low fitness load for the 2 single-insertion transgenic mosquito lines VD35 and VD26 and no load for the double-insertion transgenic mosquito line VD9. However, in cage experiments, where each of the 3 homozygous transgenic mosquitoes was mixed with nontransgenic mosquitoes, transgene frequency of all 3 lines decreased with time. Further experiments suggested that reduction of transgene frequency is a consequence of reduced mating success, reduced reproductive capacity, and/or insertional mutagenesis, rather than expression of the transgene itself. Thus, for transgenic mosquitoes released in the field to be effective in reducing malaria transmission, a driving mechanism will be required.

Myostatin gene knockdown through lentiviral-mediated delivery of shRNA for in vitro production of transgenic bovine embryos

Myostatin is described as a negative regulator of the skeletal muscle growth. Genetic engineering, in order to produce animals with double the muscle mass and that can transmit the characteristic to future progeny, may be useful. In this context, the present study aimed to analyse the feasibility of lentiviral-mediated delivery of short hairpin RNA (shRNA) targeting of myostatin into in vitro produced transgenic bovine embryos. Lentiviral vectors were used to deliver a transgene that expressed green fluorescent protein (GFP) and an shRNA that targeted myostatin. Vector efficiency was verified through in vitro murine myoblast (C2C12) cell morphology after inductive differentiation and by means of real-time PCR. The lentiviral vector was microinjected into the perivitellinic space of in vitro matured oocytes. Non-microinjected oocytes were used as the control. After injection, oocytes were fertilized and cultured in vitro. Blastocysts were evaluated by epifluorescence microscopy. Results demonstrated that the vector was able to inhibit myostatin mRNA in C2C12 cells, as the transducted group had a less amount of myostatin mRNA after 72 h of differentiation (p < 0.05) and had less myotube formation than the non-transduced group (p < 0.05). There was no difference in cleavage and blastocyst rates between the microinjected and control groups. After hatching, 3.07% of the embryos exhibited GFP expression, indicating that they expressed shRNA targeting myostatin. In conclusion, we demonstrate that a lentiviral vector effectively performed shRNA myostatin gene knockdown and gene delivery into in vitro produced bovine embryos. Thus, this technique can be considered a novel option for the production of transgenic embryos and double muscle mass animals.

Early Changes in Gene Expression Induced by Tobacco Smoke: Evidence for the Importance of Estrogen within Lung Tissue

Lung cancer is the leading cause of cancer deaths in the United States, surpassing breast cancer as the primary cause of cancer-related mortality in women. The goal of the present study was to identify early molecular changes in the lung induced by exposure to tobacco smoke and thus identify potential targets for chemoprevention. Female A/J mice were exposed to either tobacco smoke or HEPA-filtered air via a whole-body exposure chamber (6 h/d, 5 d/wk for 3, 8, and 20 weeks). Gene expression profiles of lung tissue from control and smoke-exposed animals were established using a 15K cDNA microarray. Cytochrome P450 1b1, a phase I enzyme involved in both the metabolism of xenobiotics and the 4-hydroxylation of 17 beta-estradiol (E(2)), was modulated to the greatest extent following smoke exposure. A panel of 10 genes were found to be differentially expressed in control and smoke-exposed lung tissues at 3, 8, and 20 weeks (P < 0.001). The interaction network of these differentially expressed genes revealed new pathways modulated by short-term smoke exposure, including estrogen metabolism. In addition, E(2) was detected within murine lung tissue by gas chromatography-coupled mass spectrometry and immunohistochemistry. Identification of the early molecular events that contribute to lung tumor formation is anticipated to lead to the development of promising targeted chemopreventive therapies. In conclusion, the presence of E2 within lung tissue when combined with the modulation of cytochrome P450 1b1 and other estrogen metabolism genes by tobacco smoke provides novel insight into a possible role for estrogens in lung cancer. Cancer Prev Res; 3(6); 707-17. (C) 2010 AACR.

Actions of octocoral and tobacco cembranoids on nicotinic receptors

Nicotinic acetylcholine receptors (AChRs) are pentameric proteins that form agonist-gated cation channels through the plasma membrane. AChR agonists and antagonists are potential candidates for the treatment of neurodegenerative diseases. Cembranoids are naturally occurring diterpenoids that contain a 14-carbon ring. These diterpenoids interact with AChRs in complex ways: as irreversible inhibitors at the agonist sites, as noncompetitive inhibitors, or as positive modulators, but no cembranoid was ever shown to have agonistic activity on AChRs. The cembranoid eupalmerin acetate displays positive modulation of agonist-induced currents in the muscle-type AChR and in the related gamma-aminobutyric acid (GABA) type A receptor. Moreover, cembranoids display important biological effects, many of them mediated by nicotinic receptors. Cembranoids from tobacco are neuroprotective through a nicotinic anti-apoptotic mechanism preventing excitotoxic neuronal death which in part could result from anti-inflammatory properties of cembranoids. Moreover, tobacco cembranoids also have anti-inflammatory properties which could enhance their neuroprotective properties. Cembranoids from tobacco affect nicotine-related behavior: they increase the transient initial ataxia caused by first nicotine injection into naive rats and inhibit the expression of locomotor sensitization to repeated injections of nicotine. In addition, cembranoids are known to act as anti-tumor compounds. In conclusion, cembranoids provide a promising source of lead drugs for many clinical areas, including neuroprotection, smoking-cessation, and anti-cancer therapies. (C) 2009 Elsevier Ltd. All rights reserved.

Mercury content of tobacco products

Mercury contamination of food products results from contact with soil, water, and air polluted with mercury from industrial sources, as well as from the use of mercury-containing pesticides and fungicides on plants. A method was developed for extracting mercury from tobacco products. The mercury content of various tobacco products was determined, using an atomic absorption spectrophotometer.

Transgenic lettuce seedlings carrying hepatitis B virus antigen HBsAg

The obtainment of transgenic edible plants carrying recombinant antigens is a desired issue in search for economic alternatives viewing vaccine production. Here we report a strategy for genetic transformation of lettuce plants (Lactuca sativa L.) using the surface antigen HBsAg of hepatitis B virus. Transgenic lettuce seedlings were obtained through the application of a regulated balance of plant growth regulators. Genetic transformation process was acquired by cocultivation of cotyledons with Agrobacterium tumefaciens harboring the recombinant plasmid. It is the first description of a lettuce Brazilian variety Vitória de Verão genetically modified.

Effect of respiratory muscle training on pulmonary function in preoperative preparation of tobacco smokers

OBJETIVO: Avaliar o efeito da utilização de um programa de treinamento específico dos músculos respiratórios sobre a função pulmonar em indivíduos tabagistas. MÉTODOS: Foram estudados 50 indivíduos tabagistas assintomáticos com idade superior a 30 anos, nos seguintes momentos: A0 - avaliação inicial seguida do protocolo de exercícios respiratórios; A1 - reavaliação após 10 minutos da aplicação do protocolo; e A2 -reavaliação final após duas semanas de treinamento utilizando o mesmo protocolo três vezes por semana. A avaliação foi realizada através das medidas de pressões respiratórias máximas (PImax. e PEmax.), picos de fluxo respiratórios (PFI e PFE), ventilação voluntária máxima (VVM), capacidade vital Forçada (CVF) e Volume expiratório forçado no primeiro segundo (VEF1). RESULTADOS: Não houve melhora na CVF e VEF1 da avaliação inicial para a final. Houve aumento significativo das variáveis PFI, PFE, VVM e PImax nas avaliações A1 e A2. A variável PEmax. aumentou somente na avaliação A2. CONCLUSÃO: A aplicação de protocolo de exercícios respiratórios com e sem carga adicional em indivíduos tabagistas produziu melhora imediata na performance dos músculos respiratórios, mas esta melhora foi mais acentuada após duas semanas de exercício.

Tobacco smoke-induced left ventricular remodelling is not associated with metalloproteinase-2 or -9 activation

Aim: To investigate the role of MMP-2 and MMP-9 in cardiac remodelling induced by tobacco smoke exposure in rats.Methods: Rats were allocated into two groups: C (n = 9): control animals; ETS (n = 9): exposed to tobacco smoke. After 4months, the animals underwent echocardiography, morphometric study and determination of MMP-2 and MMP-9 activity.Results: ETS rats had larger diastolic (C= 15.6 +/- 1.2 mm/kg, ETS = 18.0 +/- 0.9 mm/kg; p < 0.001) and systolic (C= 7.3 +/- 1.2 mm/kg, ETS = 9.2 0.9 mm/kg; p = 0.001) ventricular diameters adjusted for body weight. Fractional shortening (C= 53 +/- 4.8%, ETS = 48 +/- 3.3%; p = 0.031) and ejection fraction (C= 0. 89 +/- 0.03 5 ETS = 0. 86 +/- 0.02; p = 0.03 0) were smaller in the ETS group. Myocyte cross-sectional area (C= 245 8 mu m(2), ETS=253 8 mu m(2); p = 0.028) was higher in ETS rats. There were no differences in MNtP-2 (C=50 +/- 14%; ETS 43 +/- 11%, p 0.22 +/- 8) or MMP-9 (C=0.36 +/- 0.3%; ETS=0.62 +/- 0.3%, p=0.630) activity between the groups.Conclusion: MMP-2 and MMP-9 did not participate in the remodelling process induced by tobacco smoke exposure. (c) 2007 European Society of Cardiology. Published by Elsevier B.V. All rights reserved.