A Novel Stress-Induced Sugarcane Gene Confers Tolerance to Drought, Salt and Oxidative Stress in Transgenic Tobacco Plants

Background: Drought is a major abiotic stress that affects crop productivity worldwide. Sugarcane can withstand periods of water scarcity during the final stage of culm maturation, during which sucrose accumulation occurs. Meanwhile, prolonged periods of drought can cause severe plant losses. Methodology/Principal Findings: In a previous study, we evaluated the transcriptome of drought-stressed plants to better understand sugarcane responses to drought. Among the up-regulated genes was Scdr1 (sugarcane drought-responsive 1). The aim of the research reported here was to characterize this gene. Scdr1 encodes a putative protein containing 248 amino acids with a large number of proline (19%) and cysteine (13%) residues. Phylogenetic analysis showed that ScDR1is in a clade with homologs from other monocotyledonous plants, separate from those of dicotyledonous plants. The expression of Scdr1 in different varieties of sugarcane plants has not shown a clear association with drought tolerance. Conclusions/Significance: The overexpression of Scdr1 in transgenic tobacco plants increased their tolerance to drought, salinity and oxidative stress, as demonstrated by increased photosynthesis, water content, biomass, germination rate, chlorophyll content and reduced accumulation of ROS. Physiological parameters, such as transpiration rate (E), net photosynthesis (A), stomatal conductance (gs) and internal leaf CO2 concentration, were less affected by abiotic stresses in transgenic Scdr1 plants compared with wild-type plants. Overall, our results indicated that Scdr1 conferred tolerance to multiple abiotic stresses, highlighting the potential of this gene for biotechnological applications.

Gender parity and drug use: are girls catching up with boys?

Objective: To evaluate the association between gender and use of alcohol, tobacco, and other drugs in adolescents aged 10 to 18 years in the municipalities of Jacare and Diadema, Sao Paulo, Brazil. Methods: A total of 971 adolescents completed the Drug Use Screening Inventory (DUSI). Results: In our sample, 55% of adolescents were male, 33.8% reported having made use in the previous month of alcohol, 13.5% of cigarettes, and 6.4% of illicit drugs. There was no significant difference between genders in the use of alcohol, tobacco, and illicit drugs in any of the analysis (p > 0.05). The use of alcohol, tobacco, and illicit drugs was associated with the city, age, educational level, school failure, and relationship with parents (p < 0.05). Conclusions: Substance abuse among adolescents in our sample seems to follow the recent global trend towards the equalization of drug use between genders. This result should be taken into account by public health professionals in developing policies for this problem. (C) 2012 Elsevier Editora Ltda. All rights reserved.

Drug use in college students: a 13-year trend

OBJECTIVE: To analyze drug use trends among college students in 1996, 2001 and 2009. METHODS: A cross-sectional epidemiological study with a multistage stratified cluster sample with 9,974 college students was conducted in the city of Sao Paulo, southeastern Brazil. An anonymous self-administered questionnaire was used to collect information on drug use assessed in lifetime, the preceding 12 months and the preceding 30 days. The Bonferroni correction was used for multiple comparisons of drug use rates between surveys. RESULTS: There were changes in the lifetime use of tobacco and some other drugs (hallucinogens [6.1% to 8.8%], amphetamines [4.6% to 8.7%], and tranquilizers [5.7% to 8.2%]) from 1996 to 2009. Differences in the use of other drugs over the 12 months preceding the survey were also seen: reduced use of inhalants [9.0% to 4.8%] and increased use of amphetamines [2.4% to 4.8%]. There was a reduction in alcohol [72.9% to 62.1%], tobacco [21.3% to 17.2%] and marijuana [15.0% to 11.5%] use and an increase in amphetamine use [1.9% to 3.3%] in the preceeding 30 days. CONCLUSIONS: Over the 13-year study period, there was an increase in lifetime use of tobacco, hallucinogens, amphetamines, and tranquilizers. There was an increase in amphetamine use and a reduction in alcohol use during the preceding 12 months. There was an increase in amphetamine use during the preceding 30 days.

Ectopic expression of a fruit phytoene synthase from Citrus paradisi Macf. promotes abiotic stress tolerance in transgenic tobacco

Abscisic acid (ABA) is an important regulator of plant responses to environmental stresses and an absolute requirement for stress tolerance. Recently, a third phytoene synthase (PSY3) gene paralog was identified in monocots and demonstrated to play a specialized role in stress-induced ABA formation, thus suggesting that the first committed step in carotenogenesis is a key limiting step in ABA biosynthesis. To examine whether the ectopic expression of PSY, other than PSY3, would similarly affect ABA level and stress tolerance, we have produced transgenic tobacco containing a fruit-specific PSY (CpPSY) of grapefruit (Citrus paradisi Macf.). The transgenic plants contained a single- or double-locus insertion and expressed CpPSY at varying transcript levels. In comparison with the wild-type plants, the CpPSY expressing transgenic plants showed a significant increase on root length and shoot biomass under PEG-, NaCl- and mannitol-induced osmotic stress. The enhanced stress tolerance of transgenic plants was correlated with the increased endogenous ABA level and expression of stress-responsive genes, which in turn was correlated with the CpPSY copy number and expression level in different transgenic lines. Collectively, these results provide further evidence that PSY is a key enzyme regulating ABA biosynthesis and that the altered expression of other PSYs in transgenic plants may provide a similar function to that of the monocot's PSY3 in ABA biosynthesis and stress tolerance. The results also pave the way for further use of CpPSY, as well as other PSYs, as potential candidate genes for engineering tolerance to drought and salt stress in crop plants.

[Smokeless tobacco: a new risk factor for oral health? A review]

Due to migration of different ethnic groups, mainly from Africa and Asia, into Switzerland, many traditional products and habits that have not been seen before were also introduced. Smokeless tobacco (ST)--as one of those habits--is a product of increased use in Switzerland, although no sound epidemiological data are presently available. Numerous studies from North-America, Sweden, Asia and Africa could verify the carcinogenic potency of smokeless tobacco and its effects in the developement of oral cancer and different systemic disorders. For dental professionals and their team it is important to detect ST-associated lesions, and to tell the patients about the potential harmful sequelae of ST products. The present review focusses on the different types of RT products and their effects on oral health.

What Ethanol metabolites as biological markers tell us about alcohol use

Alcohol and tobacco related disorders are the two leading and most expensive causes of illness in central Europe. In addition to self reports and questionnaires, biomarkers are of relevance in diagnosis and therapy of alcohol use disorders.Traditional biomarkers such as gamma glutamyl transpeptidase or mean corpuscualr volume are indirect biomarkers and are subject to influence of age, gender and non alcohol related diseases, among others.Direct ethanol metabolites such as ethyl glucuronide (EtG), ethyl sulphate (EtS) and phosphatidylethanol (PEth) are direct metabolites of ethanol, that are positive after intake of ethyl alcohol. They represent useful diagnostic tools for identifying alcohol use even more accurately than traditional biomarkers. Each of these drinking indicators remains positive in serum and urine for a characteristic time spectrum after the cessation of ethanol intake-EtG and EtS in urine up to 7 days, EtG in hair for months after ethanol has left the body. Applications include clinical routine use, emergency room settings, proof of abstinence in alcohol rehabilitation programs, driving under influence offenders, workplace testing, assessment of alcohol intake in the context of liver transplantation and fetal alcohol syndrome.

Childhood sexual victimization: Risk factor for drug use and sexual risk behaviors

The purpose of this dissertation was to survey men in the Harris County Jail (HCJ) to establish a more valid estimate of childhood sexual abuse (CSA) prevalence in a jailed-based population; to assess whether inmates with a history of CSA were at greater risk for use of drugs and alcohol and engaging in high-risk sexual behaviors than those without histories of childhood sexual abuse. ^ The first study determined the prevalence of childhood sexual abuse among incarcerated males in a county jail. In this study, sixty-three percent of the subjects reported having been sexually abused. Sixty-one percent reported abuse pre-puberty and 10% reported abuse post puberty. In pre-puberty abuse the initiation of first abuse occurred at a mean age of 5.6 years (SD 5.096, range: 2–13 years). ^ The second study explored the association between inmates with histories of CSA as a risk factor for sexual risk behaviors. A history of sexual abuse did not appear to be associated with an elevated risk of sexual risk behaviors. ^ The third study explored a history of drug use and a history of CSA among the inmates. A chi-square test showed that the inmates who reported a history of CSA, was significantly greater for the following drugs: Marijuana (02), Crack (03), Heroin/Morphine (.03), Amphetamines/Speed (01), Downers/Barbiturates (.001), Methamphetamine/Crystal Meth (.001), Valium .02), LSD/Acid (.001), and Inhalants (.001), p < .05). Significance was not found in alcohol, tobacco, cocaine, Quaaludes and methadone. ^ The research from this study provides empirical data supporting previous research. The current data shows that incarcerated inmates have a high prevalence of childhood sexual abuse and drug use. Sexual victimization as a child does not appear to be associated with an elevated risk of unsafe sexual behaviors. However, men who used drugs were twice as likely to have engaged in unprotected sex with casual and regular partners, and rarely used condoms with paid sex. Although our study methods do not permit a causal explanation for this association, we believe it is of concern. Finally, data in this study shows that sexually abused children are likely candidates for adult criminal behavior. ^

Investigating the epidemic of heroin use in Dar es Salaam, Tanzania: Recognizing the issues of heroin smokers

Since heroin was introduced to East Africa during the 1980s, heroin use practices have changed rapidly in response to various internal and external pressures. The aim of this study was to identify and describe the population of heroin users and locations of heroin use in Dar es Salaam, Tanzania, in order to understand recent contexts of heroin use. The study took place between June 30 and August 19, 2011, in all three districts (Kinondoni, Ilala, and Temeke) of Dar es Salaam. We mapped sites using a Global Positioning System device, counted numbers of heroin users, and conducted informal interviews with heroin users. The mixed-methods analyses of the data included quantifying the basic demographic and aggregate information about the sites and heroin users, as well as qualitative analysis and coding of fieldnotes from observations and responses to interviews which was used to identify themes and characteristics of heroin users. ^ We identified a total of 150 sites and counted a total of 1046 male and 46 female non-injecting drug users and 78 male and 9 female injecting drug users (IDUs) of heroin. We found that social organization existed at some of the sites, with 31% (n=47) of sites reporting having a leader and 44% (n=66) of sites reporting mutual aid between users frequenting the site. We had difficulty locating IDUs and female drug users, and the majority of users we encountered were heroin smokers of kokteli, a mixture of heroin, cannabis, and/or tobacco which is smoked like a cigarette. ^ This research highlighted heroin smokers’ desire for access to drug treatment services. The current methadone-based medication assisted treatment (MAT) program is funded and operates as an HIV prevention program for IDUs to reduce HIV infection in this population and slow or stop the spread of a second wave of HIV infection in the general population. However, smokers perceived MAT to be primarily a drug use prevention or cessation program and felt unjustly neglected from the intervention, leading to a tense relationship with IDUs. From a public health standpoint, future interventions should include heroin smokers to prevent HIV transmission. ^

Developmental changes in DNA methylation of the two tobacco pollen nuclei during maturation

Changes in DNA methylation during tobacco pollen development have been studied by confocal fluorescence microscopy using a monoclonal anti-5-methylcytosine (anti-m5C) antibody and a polyclonal anti-histone H1 (anti-histone) antibody as an internal standard. The specificity of the anti-m5C antibody was demonstrated by a titration series against both single-stranded DNA and double-stranded DNA substrates in either the methylated or unmethylated forms. The antibody was found to show similar kinetics against both double- and single-stranded DNA, and the fluorescence was proportional to the amount of DNA used. No signal was observed with unmethylated substrates. The extent of methylation of the two pollen nuclei remained approximately constant after the mitotic division that gave rise to the vegetative and generative nuclei. However, during the subsequent development of the pollen, the staining of the generative nucleus decreased until it reached a normalized value of \documentclass[12pt]{minimal} \usepackage{wasysym} \usepackage{amsmath} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \setlength{\oddsidemargin}{-69pt} \begin{document} \begin{equation*}\frac{1}{5}\end{equation*}\end{document} of that of the vegetative nucleus. The use of a confocal microscope makes these data independent of possible focusing artefacts. The anti-histone antibody was used as a control to show that, while the antibody staining directed against 5-methylcytosine changed dramatically during pollen maturation, the histone signal did not. We observed the existence of structural dimorphism amongst tobacco pollen grains, the majority having three pollen apertures and the rest with four. However, the methylation changes observed occurred to the same extent in both subclasses.

Use of Ubiquitin Fusions to Augment Protein Expression in Transgenic Plants1

A major goal of plant biotechnology is the production of genetically engineered crops that express natural or foreign proteins at high levels. To enhance protein accumulation in transgenic plants, we developed a set of vectors that express proteins and peptides as C-terminal translational fusions with ubiquitin (UBQ). Studies of several proteins in tobacco (Nicotiana tabacum) showed that: (a) proteins can be readily expressed in plants as UBQ fusions; (b) by the action of endogenous UBQ-specific proteases (Ubps), these fusions are rapidly and precisely processed in vivo to release the fused protein moieties in free forms; (c) the synthesis of a protein as a UBQ fusion can significantly augment its accumulation; (d) proper processing and localization of a protein targeted to either the apoplast or the chloroplast is not affected by the N-terminal UBQ sequence; and (e) single amino acid substitutions surrounding the cleavage site can inhibit in vivo processing of the fusion by Ubps. Noncleavable UBQ fusions of β-glucuronidase became extensively modified, with additional UBQs in planta. Because multiubiquitinated proteins are the preferred substrates of the 26S proteasome, noncleavable fusions may be useful for decreasing protein half-life. Based on their ability to augment protein accumulation and the sequence specificity of Ubps, UBQ fusions offer a versatile way to express plant proteins.

Plasma Membrane-Associated Actin in Bright Yellow 2 Tobacco Cells1 : Evidence for Interaction with Microtubules

Plasma membrane ghosts form when plant protoplasts attached to a substrate are lysed to leave a small patch of plasma membrane. We have identified several factors, including the use of a mildly acidic actin stabilization buffer and the inclusion of glutaraldehyde in the fixative, that allow immunofluorescent visualization of extensive cortical actin arrays retained on membrane ghosts made from tobacco (Nicotiana tabacum L.) suspension-cultured cells (line Bright Yellow 2). Normal microtubule arrays were also retained using these conditions. Membrane-associated actin is random; it exhibits only limited coalignment with the microtubules, and microtubule depolymerization in whole cells before wall digestion and ghost formation has little effect on actin retention. Actin and microtubules also exhibit different sensitivities to the pH and K+ and Ca2+ concentrations of the lysis buffer. There is, however, strong evidence for interactions between actin and the microtubules at or near the plasma membrane, because both ghosts and protoplasts prepared from taxol-pretreated cells have microtubules arranged in parallel arrays and an increased amount of actin coaligned with the microtubules. These experiments suggest that the organization of the cortical actin arrays may be dependent on the localization and organization of the microtubules.

Heat-Shock-Induced Changes in Intracellular Ca2+ Level in Tobacco Seedlings in Relation to Thermotolerance1

Exposure of plants to elevated temperatures results in a complex set of changes in gene expression that induce thermotolerance and improve cellular survival to subsequent stress. Pretreatment of young tobacco (Nicotiana plumbaginifolia) seedlings with Ca2+ or ethylene glycol-bis(β-aminoethylether)-N,N,N′,N′-tetraacetic acid enhanced or diminished subsequent thermotolerance, respectively, compared with untreated seedlings, suggesting a possible involvement of cytosolic Ca2+ in heat-shock (HS) signal transduction. Using tobacco seedlings transformed with the Ca2+-sensitive, luminescent protein aequorin, we observed that HS temperatures induced prolonged but transient increases in cytoplasmic but not chloroplastic Ca2+. A single HS initiated a refractory period in which additional HS signals failed to increase cytosolic Ca2+. However, throughout this refractory period, seedlings responded to mechanical stimulation or cold shock with cytosolic Ca2+ increases similar to untreated controls. These observations suggest that there may be specific pools of cytosolic Ca2+ mobilized by heat treatments or that the refractory period results from a temporary block in HS perception or transduction. Use of inhibitors suggests that HS mobilizes cytosolic Ca2+ from both intracellular and extracellular sources.

Use of Arabidopsis thaliana defense-related mutants to dissect the plant response to pathogens.

The plant defense response to microbial pathogens had been studied primarily by using biochemical and physiological techniques. Recently, several laboratories have developed a variety of pathosystems utilizing Arabidopsis thaliana as a model host so that genetic analysis could also be used to study plant defense responses. Utilizing a pathosystem that involves the infection of Arabidopsis with pathogenic pseudomonads, we have cloned the Arabidopsis disease-resistance gene RPS2, which corresponds to the avirulence gene avrRpt2 in a gene-for-gene relationship. RPS2 encodes a 105-kDa protein containing a leucine zipper, a nucleotide binding site, and 14 imperfect leucine-rich repeats. The RPS2 protein is remarkably similar to the product of the tobacco N gene, which confers resistance to tobacco mosaic virus. We have also isolated a series of Arabidopsis mutants that synthesize decreased levels of an Arabidopsis phytoalexin called camalexin. Analysis of these mutants indicated that camalexin does not play a significant role in limiting growth of avirulent Pseudomonas syringae strains during the hypersensitive defense response but that it may play a role in limiting the growth of virulent strains. More generally, we have shown that we can utilize Arabidopsis to systematically dissect the defense response by isolation and characterization of appropriate defense-related mutants.

Catalytic effect of MCM-41 in the pyrolysis of tobacco and several substances used as additives in the preparation of commercial cigarettes

Comunicación presentada en forma de póster en el "12th Mediterranean Congress of Chemical Engineering", Barcelona (Spain), November 15-18, 2011.

TGA/FTIR study of the MCM-41-catalytic pyrolysis of tobacco and tobacco–glycerol mixtures

In this work, the catalytic effect of MCM-41 on the qualitative composition of the gases evolved in the pyrolysis of glycerol, tobacco and tobacco–glycerol mixtures has been studied by TGA/FTIR. The results obtained reflect that the amount of volatile products obtained from tobacco or from tobacco containing mixtures is noticeably decreased if the catalyst is used as a tobacco additive. The addition of the catalyst also produces noticeable changes in the composition of the gases obtained at each temperature. Such changes are more significant in the case of the pyrolysis of the tobacco–glycerol–mixture than in the pyrolysis of tobacco. The evolution patterns for different types of compounds reveal that the catalyst contributes to a noticeable decrease of the yields of methane, CO, CO2 and carbonylic compounds, which could be interesting from the point of view of the use of MCM-41 as a tobacco additive in order to reduce tobacco smoke toxicity.