Transgene Silencing and Transgene-Derived siRNA Production in Tobacco Plants Homozygous for an Introduced AtMYB90 Construct

Transgenic tobacco (Nicotiana tabacum) lines were engineered to ectopically over-express AtMYB90 (PAP2), an R2–R3 Myb gene associated with regulation of anthocyanin production in Arabidopsis thaliana. Independently transformed transgenic lines, Myb27 and Myb237, accumulated large quantities of anthocyanin, generating a dark purple phenotype in nearly all tissues. After self-fertilization, some progeny of the Myb27 line displayed an unexpected pigmentation pattern, with most leaves displaying large sectors of dramatically reduced anthocyanin production. The green-sectored 27Hmo plants were all found to be homozygous for the transgene and, despite a doubled transgene dosage, to have reduced levels of AtMYB90 mRNA. The observed reduction in anthocyanin pigmentation and AtMYB90 mRNA was phenotypically identical to the patterns seen in leaves systemically silenced for the AtMYB90 transgene, and was associated with the presence of AtMYB90-derived siRNA homologous to both strands of a portion of the AtMYB90 transcribed region. Activation of transgene silencing in the Myb27 line was triggered when the 35S::AtMYB90 transgene dosage was doubled, in both Myb27 homozygotes, and in plants containing one copy of each of the independently segregating Myb27 and Myb237 transgene loci. Mapping of sequenced siRNA molecules to the Myb27 TDNA (including flanking tobacco sequences) indicated that the 3′ half of the AtMYB90 transcript is the primary target for siRNA associated silencing in both homozygous Myb27 plants and in systemically silenced tissues. The transgene within the Myb27 line was found to consist of a single, fully intact, copy of the AtMYB90 construct. Silencing appears to initiate in response to elevated levels of transgene mRNA (or an aberrant product thereof) present within a subset of leaf cells, followed by spread of the resulting small RNA to adjacent leaf tissues and subsequent amplification of siRNA production

Etude du comportement agronomique de tabacs industriels transgeniques présentant une activité nitrate réductase élevée

National audienc

A Eukaryotic Translation Initiation Factor 4E (eIF4E) is responsible for the “va” tobacco recessive resistance to potyviruses

Potato virus Y (PVY), the type member of the genus Potyvirus, is transmitted by aphids and can cause severe damage in several solanaceous family crops. In Nicotiana tabacum, a large genome deletion conferring resistance to PVY, the va gene, is commonly used. This resistance is unfortunately associated with lower tobacco quality parameters, potentially due to the presence of several other important genes in the deleted region. In the present study, we have used next-generation sequencing to analyze the transcriptome of a dozen of tobacco F7 recombinant inbred lines (RILs) segregating for PVY resistance. After comparison with a reference transcriptome, genes differentially expressed between resistant and susceptible plants were identified. About 30 candidate sequences were selected, including a sequence annotated as encoding an eukaryotic translation initiation factor 4E (eIF4E), which was strongly expressed in susceptible plants but not in resistant ones. Other differentially expressed candidates are mostly related to photosynthesis. A complete correlation between susceptibility and expression of this eIF4E sequence was confirmed by amplification in 91 F8 RILs and in a segregating F2 population. The gene was mapped on chromosome 21 of the tobacco genetic map and corresponds to an eIF4E isoform derived from the N. sylvestris parent of tobacco. Final confirmation of the identification of the va gene came from the analysis of two tobacco lines with missense mutations in the eIF4E gene and which correspondingly showed resistance to PVY infection. Screening of a large collection of tobacco accessions revealed a strong correlation between the status of this eIF4 gene and PVY resistance, but the identification of a few resistant accessions with an apparently intact gene suggests the possible existence of alternative resistance sources. The identification of the va gene and of molecular markers linked to it or to the large deletion associated with it opens the way to breeding efforts aimed at breaking the linkage drag associated with this valuable resistance gene

Potato virus Y resistance in tobacco: tolerance to viral necrosis as an alternative strategy against PVY resistant-breaking variants

[i]Potato virus Y[/i] resistance in tobacco: tolerance to viral necrosis as an alternative strategy against PVY resistant-breaking variants. 16. Rencontres de Virologie Végétale (RVV 2017

Differential impact of retrotransposon populations on the genome of allotetraploid tobacco (Nicotiana tabacum)

International audienceLTR-retrotransposons contribute substantially to the structural diversity of plant genomes. Recent models of genome evolution suggest that retrotransposon amplification is offset by removal of retrotransposon sequences, leading to a turnover of retrotransposon populations. While bursts of amplification have been documented, it is not known whether removal of retrotransposon sequences occurs continuously, or is triggered by specific stimuli over short evolutionary periods. In this work, we have characterized the evolutionary dynamics of four populations of copia-type retrotransposons in allotetraploid tobacco (Nicotiana tabacum) and its two diploid progenitors Nicotiana sylvestris and Nicotiana tomentosiformis. We have used SSAP (Sequence-Specific Amplification Polymorphism) to evaluate the contribution retrotransposons have made to the diversity of tobacco and its diploid progenitor species, to quantify the contribution each diploid progenitor has made to tobacco's retrotransposon populations, and to estimate losses or amplifications of retrotransposon sequences subsequent to tobacco's formation. Our results show that the tobacco genome derives from a turnover of retrotransposon sequences with removals concomitant with new insertions. We have detected unique behaviour specific to each retrotransposon population, with differences likely reflecting distinct evolutionary histories and activities of particular elements. Our results indicate that the retrotransposon content of a given plant species is strongly influenced by the host evolutionary history, with periods of rapid turnover of retrotransposon sequences stimulated by allopolyploidy

Bronquiectasias : tratamento cirúrgico : análise de 437 casos

-Introdução: Este trabalho apresenta a experiência cirúrgica em 437 pacientes submetidos à ressecção pulmonar para tratamento de bronquiectasias, avaliando evolução pós-operatória e o impacto quanto à melhora dos sintomas. -Métodos: Foram analisados retrospectivamente os prontuários médicos de 437 pacientes submetidos à ressecção pulmonar para tratamento de bronquiectasias em uma única instituição, entre Janeiro de 1978 e Dezembro de 2010. Os pacientes tinham 16 ou mais anos de idade, e as bronquiectasias foram diagnosticadas pelas manifestações clínicas e por tomografia computadorizada do tórax. Pacientes portadores de fibrose cística e os submetidos a transplante pulmonar foram excluídos. -Resultados: A amostra incluiu 437 pacientes (249 do sexo feminino; 188 do masculino). A idade média dos pacientes na época da cirurgia era de 38,45 anos (variando de 16 a 80 anos). O sintoma mais comum na apresentação era tosse produtiva (79,4%). A causa identificada como mais comum das bronquiectasias foi infecção respiratória na infância (49,2%). Cirurgia unilateral foi realizada em 415 pacientes (95,0%), e em 344 toda a área com bronquiectasias foi ressecada. Dentre os procedimentos cirúrgicos utilizados, o principal foi lobectomia média em 147 pacientes (33,6%) e segmentectomia (32,3%). Após a cirurgia, 94,4% de 267 pacientes questionados referiram melhora de seus sintomas, 68,9% deles considerando-se assintomáticos. Ocorreu 1 (um) óbito no pós operatório. -Conclusões: O tratamento cirúrgico das bronquiectasias mostrou-se altamente eficiente, em termos de melhora dos sintomas dos indivíduos acometidos, sendo realizado com taxas de morbidade aceitáveis, mortalidade praticamente nula, e com excelentes resultados em termos de melhora dos sintomas.Background. This work presents the surgical experience of 437 patients undergoing to lung resection for the treatment of bronchiectasis, evaluating both the impact on postoperative clinical improvement and complications of the surgical procedure. Methods It was retrospectively analyzed the medical charts of 437 consecutive patients who underwent a surgical lung resection for treatment of bronchiectasis in a single institution, between January 1978 and December 2010. Patients were 16 years or older and bronchiectasis was diagnosed by symptoms and computed tomography. Patients with cystic fibrosis and bronchiectasis the ones that underwent to lung transplantation were excluded. Results The study sample included 437 patients (249 females; 188 males). Mean age at time of surgery was 38.45 years old (range: 16 to 80). The most common presenting symptoms were cough (79.4%). The most common etiology of bronchiectais was childhood infections (49.2%). Unilateral approach was performed in 415 (95.0%) and in 344 the total bronchiectasic site were resected. Several surgical procedures were used, the majority was medium lobectomy in 147 (33.6%) and segmentectomy (32.3%). After surgery, 94.4% of the patients improved their symptoms, 68,9% following practically assymptomatic. One postoperative death occurred. Conclusions Bronchiectasis may be treated by surgical management, particularly in developing countries where it is very prevalent. It can be performed with acceptable morbidity rates, practically without mortality, and with excelent results in terms of symptoms improvement