RESUMEN

Prolyl 4-hydroxylase (P4H) generates hydroxyproline residues in proteins. Two classes of P4H have been found in plants. Type 1 P4H has a signal anchor at the N-terminus, while type 2 P4H has both an N-terminal signal peptide and a C-terminal toxin homology domain (Tox1 domain) with six conserved cysteine residues. We analyzed the localization of tobacco type 2 P4H (NtP4H2.2) in tobacco BY-2 cells. Cell fractionation studies, immunostaining of cells, and GFP fusion study indicated that NtP4H2.2 localizes predominantly to the Golgi apparatus and is a peripheral membrane protein associated with the luminal side of organelles. Expression of the GFP-Tox1 domains of NtP4H2.2 and another tobacco type 2 P4H NtP4H2.1 in BY-2 cells and Arabidopsis epidermal cells indicated that these proteins were targeted to the Golgi. The Tox1 domains from Arabidopsis and rice type 2 P4Hs also directed GFP to the Golgi in tobacco BY-2 cells. The Tox1 domain of NtP4H2.2 increased the membrane association of GFP, and mutation of the cysteine residues in this domain abolished Golgi localization. Furthermore, the catalytic domain of NtP4H2.2 also directed GFP to the Golgi. Thus, the Tox1 domains of plant P4Hs are the Golgi localization domains, and tobacco P4H2.2 localizes to the Golgi by the action of both this domain and the catalytic domain.

Asunto(s)

Arabidopsis , Aparato de Golgi , Nicotiana , Dominios Proteicos , Aparato de Golgi/metabolismo , Nicotiana/metabolismo , Arabidopsis/enzimología , Arabidopsis/metabolismo , Prolil Hidroxilasas/metabolismo , Proteínas de Plantas/metabolismo , Proteínas de Plantas/química , Proteínas de Plantas/genética , Transporte de Proteínas , Secuencia de Aminoácidos

RESUMEN

During the self-assembly of π-conjugated molecules, linkers and substituents can potentially add supportive noncovalent intermolecular interactions to π-stacking interactions. Here, we report the self-assembly behavior of thienopyrrole-fused thiadiazole (TPT) fluorescent dyes that possess ester or ether linkers and dodecyloxy side chains in solution and the condensed phase. A comparison of the self-association behavior of the ester- and ether-bridged compounds in solution using detailed UV-vis, fluorescence, and NMR spectroscopic studies revealed that the subtle replacement of the ether linkers by ester linkers leads to a distinct increase in the association constant (ca. 3-4 fold) and the enthalpic contribution (ca. 3 kcal mol-1 ). Theoretical calculations suggest that the ester linkers, which are in close proximity to one another due to the π-stacking interactions, induce attractive electrostatic forces and augment self-association. The self-assembly of TPT dyes into well-defined 1D clusters with high aspect ratios was observed, and their morphologies and crystallinity were investigated using SEM and X-ray diffraction analyses. TPTs with ester linkers exhibit a columnar liquid crystalline mesophase in the condensed phase.

Asunto(s)

Tiadiazoles , Ésteres , Éter , Éteres , Pirroles , Electricidad Estática

RESUMEN

Tomato (Solanum lycopersicum L.) plants were transformed with an antisense construct of a cDNA encoding tomato telomere-binding protein (LeTBP1) to describe the role of a telomere-binding protein at the whole plant level. Fruit size decreased corresponding to the degree of suppression of LeTBP1 expression. This inhibition of fruit development was likely due to a decrease in the number of seeds in the LeTBP1 antisense plants. Pollen fertility and pollen germination rate decreased in accordance with the degree of suppression of LeTBP1 expression. Ovule viability was also reduced in the LeTBP1 antisense plants. Although plant height was somewhat reduced in the antisense plants compared to the control plants, the number and weight of leaves were unaffected by LeTBP1 suppression. The number and morphology of flowers were also normal in the antisense plants. These indicate that reduced fertility in the antisense plants is not an indirect effect of altered vegetative growth. LeTBP1 expression was sensitive to temperature stress in wild-type plants. We conclude that LeTBP1 plays a critical role in seed and fruit development rather than vegetative growth and flower formation.

Asunto(s)

Frutas/crecimiento & desarrollo , Semillas/crecimiento & desarrollo , Solanum lycopersicum/fisiología , Supresión Genética/genética , Proteínas de Unión a Telómeros/genética , Cromosomas de las Plantas/genética , ADN sin Sentido/genética , ADN Complementario/genética , ADN de Plantas/genética , Flores/genética , Flores/crecimiento & desarrollo , Flores/fisiología , Frutas/genética , Frutas/fisiología , Expresión Génica/genética , Regulación de la Expresión Génica de las Plantas/genética , Solanum lycopersicum/citología , Solanum lycopersicum/genética , Solanum lycopersicum/crecimiento & desarrollo , Meristema/citología , Óvulo Vegetal/fisiología , Hojas de la Planta/genética , Hojas de la Planta/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Modificadas Genéticamente , Polen/fisiología , Semillas/genética , Semillas/fisiología , Estrés Fisiológico , Proteínas de Unión a Telómeros/metabolismo , Temperatura

RESUMEN

Plant specific O-glycosylation of proteins includes the attachment of arabinogalactan to hydroxyproline (Hyp) residues. These Hyp residues are generated from peptidyl proline residues by the action of prolyl 4-hydroxylase which requires the ferrous ion. We investigated the effect of the ferrous chelator, 2,2'-dipyridyl on tobacco plants, and found that such treatment reduced the arabinogalactosylation of proteins.

Asunto(s)

2,2'-Dipiridil/farmacología , Galactanos/metabolismo , Quelantes del Hierro/farmacología , Nicotiana/efectos de los fármacos , Nicotiana/metabolismo , Glicosilación/efectos de los fármacos , Especificidad de la Especie

RESUMEN

Auxin transport network, which is important in the integration of plant developmental signals, depends on differential expression of the auxin efflux carrier PIN gene family. We cloned three tomato PIN (referred as SlPIN) cDNAs and examined their expression patterns in fruit and other organs. The expression of SlPIN1 and SlPIN2 was highest in very young fruit immediately after anthesis, whereas the expression of SlPIN3 was low at this same stage of fruit development. SlPIN2::GUS was expressed in ovules at anthesis and in young developing seeds at 4 days after anthesis, while SlPIN1::GUS was expressed in whole fruit. The DR5::GUS auxin-responsive reporter gene was expressed in the fruit and peduncle at anthesis and was higher in the peduncle 4 days after anthesis. These studies suggest that auxin is likely transported from young seeds by SlPIN1 and SlPIN2 and accumulated in peduncles where SlPIN gene expression is low in tomato. The possible role of SlPINs in fruit set was discussed.

Asunto(s)

Proteínas Portadoras/genética , Ácidos Indolacéticos/metabolismo , Solanum lycopersicum/metabolismo , Secuencia de Aminoácidos , Secuencia de Bases , Proteínas Portadoras/química , Proteínas Portadoras/metabolismo , Clonación Molecular , Cartilla de ADN , ADN Complementario , Genes de Plantas , Genes Reporteros , Solanum lycopersicum/genética , Solanum lycopersicum/crecimiento & desarrollo , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Regiones Promotoras Genéticas , Homología de Secuencia de Aminoácido

RESUMEN

The enzyme NAD-dependent sorbitol dehydrogenase (SDH) is well characterized in the Rosaceae family of fruit trees, which synthesizes sorbitol as a translocatable photosynthate. Expressed sequence tags of SDH-like sequences have also been generated from various non-Rosaceae species that do not synthesize sorbitol as a primary photosynthetic product, but the physiological roles of the encoded proteins in non-Rosaceae plants are unknown. Therefore, we isolated an SDH-like cDNA (SDL) from tomato (Lycopersicon esculentum Mill.). Genomic Southern blot analysis suggested that SDL exists in the tomato genome as a single-copy gene. Northern blot analysis showed that SDL is ubiquitously expressed in tomato plants. Recombinant SDL protein was produced and purified for enzymatic characterization. SDL catalyzed the interconversion of sorbitol and fructose with NAD (H). SDL showed highest activity for sorbitol among the several substrates tested. SDL showed no activity with NADP+. Thus, SDL was identified as a SDH, although the Km values and substrate specificity of SDL were significantly different from those of SDH purified from the Japanese pear (Pyrus pyrifolia), a Rosaceae fruit tree. In addition, tomato was transformed with antisense SDL to evaluate the contribution of SDL to SDH activity in tomato. The transformation decreased SDH activity to approximately 50% on average. Taken together, these results provide molecular evidence of SDH in tomato, and SDL was renamed LeSDH.

Asunto(s)

L-Iditol 2-Deshidrogenasa/genética , L-Iditol 2-Deshidrogenasa/metabolismo , Solanum lycopersicum/enzimología , Secuencia de Aminoácidos , Northern Blotting , Southern Blotting , ADN sin Sentido , Fructosa/metabolismo , Regulación de la Expresión Génica de las Plantas , Solanum lycopersicum/genética , Datos de Secuencia Molecular , NAD/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Rosaceae/enzimología , Homología de Secuencia de Aminoácido , Sorbitol/metabolismo , Especificidad por Sustrato , Transformación Genética