Effects of flooding stress in tomato plants transformed with different levels of mitochondrial sHSP23 6 / Efeitos do estresse por alagamento em plantas de tomateiro transformada com diferentes níveis de sHSP23 6 mitocondrial

Abstract Soil flooding is an environmental stressor for crops that can affect physiological performance and reduce crop yields. Abiotic stressors cause changes in protein synthesis, modifying the levels of a series of proteins, especially the heat shock proteins (HSP), and these proteins can help protect the plants against abiotic stress. The objective of this study was to verify if tomato plants cv. Micro-Tom from different genotypes with varying expression levels of MT-sHSP23.6 (mitochondrial small heat shock proteins) have different responses physiological to flooding. Plants from three genotypes (untransformed, MT-sHSP23.6 sense expression levels and MT-sHSP23.6 antisense expression levels) were cultivated under controlled conditions. After 50 days, the plants were flooded for 14 days. After this period half of the plants from each genotype were allowed to recover. Chlorophyll fluorescence, gas exchange, chlorophyll index, leaf area and dry matter were evaluated. Flood stress affected the photosynthetic electron transport chain, which is related to inactivation of the oxygen-evolving complex, loss of connectivity among units in photosystem II, oxidation-reduction of the plastoquinone pool and activity of photosystem I. The genotype with MT-sHSP23.6 sense expression levels was less sensitive to stress from flooding.

Resumo O alagamento do solo é um estressor ambiental para as culturas e pode afetar o desempenho fisiológico e reduzir a produtividade das culturas. Estresses abióticos causam mudanças na síntese de proteínas, modificando os níveis de uma série de proteínas, em especial as proteínas de choque térmico (HSP) e essas proteínas são conhecidas por proteger as plantas contra estresses abióticos. O objetivo deste estudo foi verificar se as plantas do tomateiro cv. Micro-Tom de distintos genótipos com diferentes níveis de expressão da MT-sHSP23.6 (proteínas mitocondriais de choque térmico com pequena massa molecular), têm diferentes respostas fisiológicas ao alagamento. As plantas de três genótipos (não-transformado, transformado com orientação antisense e transformado com orientação sense para MT-sHSP23.6) foram cultivadas sob condições controladas. Após 50 dias as plantas foram alagadas durante 14 dias. Após esse período as plantas de cada genótipo foram recuperadas. Foram avaliados fluorescência da clorofila, trocas gasosas, índice de clorofila, área foliar e massa seca. O estresse por alagamento afetou a cadeia de transporte de elétrons da fotossíntese, que está relacionado à inativação do complexo de evolução do oxigênio, perda da conectividade entre as unidades do fotossistema II, de oxidação e redução do pool de plastoquinona e atividade do fotossistema I. O genótipo com orientação sense MT-sHSP23.6 foi menos sensível ao estresse por alagamento.

Studies on relationship between the expression of hTNF-alpha gene and photosynthesis in Anabaena sp. IB02 / 生物工程学报

The effects of illumination on growth of Anabaena sp. IB02 and hTNF-alpha expression were studied. Photosynthetic activity, PS I and PS II activity of Anabaena sp. IB02 were assayed. Illumination enhanced the growth of Anabaena sp. IB02 and hTNF-a expression. Some relations were observed between hTNF-alpha expression and ture photosynthesis activity, PS I, PS II activity of Anabaena sp. IB02. Significant differences of the photosynthetic activity of host were detected simultaneously when hTNF-a expressed: the respiration rate increased (-68%), the light saturation point descended (+66%), all these suggested that the metabolic charge of host were increased and grow faster than wild type under low illumination.

NaCl induced changes in photosystem stoichiometry and photosynthetic activity of the cyanobacterium Microcystis aeruginosa Kütz.

Exposure to 0.4 M NaCl resulted in higher PS I/PS II stoichiometry and increase in the rate of photosynthesis in planktonic cyanobacterium M. aeruginosa. Altered ratios of PS I/PS II as well as photosynthesis and respiration were stabilized within 72 hr of exposure to salt, leading to adaptation of the organism to the changed conditions.

Cloning and analysis of psaB cDNA of Dunaliella salina / 生物工程学报

One pair of degenerate primer was designed according to conserved motifs of the psaB (A2 subunit of photosystem I) of Chlamydomonas reinhardtii, Chlamydomonas moewusii, Chlorella vulgaris and Mesostigma viride, and a total RNA of Dunaliella salina (D. salina) was extracted with TRIzol reagent. A cDNA fragment, about 1.8kb in length, from green algal D. salina was obtained through RT-PCR method. The resulting PCR product was cloned into T-vector and screened to determine its sequence. Homologous analysis of the deduced amino acid sequence was performed by BLAST and subsequeqtly compared with GenBank data. The obtained cDNA sequence was 1815 bp long, which encodes 605 amino acids (GenBank accession number: AY820754). The sequence shared high homologue with the following psaB: Chlamydomonas reinhardtii 92%, Chlamydomonas moewusii 91%, Chlorella vulgaris 86%, Mesostigma viride 85%, Physcomitrella patens subsp. Patens 85% and Nephroselmis olivacea 84%. It can be concluded that the cloned sequence is psaB cDNA fragment from D. salina.

Addition of C-terminal histidyl tags to PsaL and PsaK1 proteins of cyanobacterial photosystem I.

In vitro mutagenesis was used to produce two photosystem I mutants of the cyanobacterium Synechocystis sp. PCC 6803. The mutant HK and HL contained hexahistidyl tags at the C-termini of the PsaK1 and PsaL subunits, respectively. The HK mutant contained wild-type amounts of trimeric PS I complexes, but the level of hexahistidine-tagged PsaK1 was found only ten per cent in the PS I complexes and membranes of the wild type level. Therefore, attachment of a tag at the C-terminus interferes with the expression or assembly of PsaK1. In contrast, the HL mutant contained a similar level of tagged PsaL as that in the wild type. However, trimeric PS I complexes could not be obtained from this strain, indicating that the C-terminus of PsaL is involved in the formation of PS I trimers. Hexahistidine-tagged complexes of the HL and HK strains could not be purified with Nickel-affinity chromatography, unless photosystem I was denatured with urea, demonstrating that tagged C-termini of PsaK1 and PsaL were embedded inside of the PS I complex. Protection of the C-terminus from trypsin cleavage further supported this conclusion. Thus, histidine tagging allowed us to demonstrate role of C-termini of two proteins of photosystem I.

Shortening of life-time of the pair [P680+Pheo-] contributes to general inhibitory effect of dinoseb on electron transfer in PS-II.

It is shown that dinoseb, added to subchloroplast photosystem-II (PS-II) preparations from pea at a concentration higher than 5 microM, along with blocking the electron transfer on the acceptor side of PS-II, induces the following effects revealing its capability to have redox interaction with the components of PS-II reaction center (RC)-pheophytin (Pheo) and chlorophyll P680: (1) acceleration of the dark relaxation of absorbance (delta A) and chlorophyll fluorescence (delta F) changes related to photoreduction of Pheo as a result of the photoreaction [P680Pheo] [symbol: see text] [P680Pheo-] that leads to elimination of the delta A and delta F at a concentration of the inhibitor higher than 50 microM; (2) lowering of the maximum level of fluorescence (F) due to a decrease of delta F under the condition when the electron acceptor, QA, is reduced; (3) loss of the described effects of dinoseb and appearance of its capability to donate electron to RC of PS-II in the presence of dithionite which reduces dinoseb in the dark; (4) inhibition of delta A related to photooxidation of P680; (5) activation of delta A related to photooxidation P700 in photosystem-I (PS-I) preparations (a similar effect is observed upon the addition of 0.2 mM methylviologen). It is suggested that redox interaction with the pair [P680+Pheo-] leading to the shortening of its life-time contributes to the general effect of inhibition of electron transfer in PS-II by dinoseb.

Analysis of the activity of promoters from two photosynthesis-related genes psaF and petH of spinach in a monocot plant, rice.

The subunit III of photosystem I and ferredoxin-NADP(+)-oxidoreductase are encoded by nuclear genes, namely psaF and petH. The activity of their promoters from spinach has been evaluated in transgenic tobacco earlier. Evaluation of the activity of these Dicotyledoneae-specific promoters has been carried out in a monocot system (i.e. rice) by transient gene expression system, based on electroporation-mediated gene delivery into protoplasts from leaves and roots. It has been found that various promoter deletions show higher activity in leaf protoplasts and elements for quantitative response are widely distributed. Transgenic rice has also been produced with a petH promoter and gus reporter gene construct. Although petH promoter is a weak promoter in comparison to the 35S promoter, it expresses well in green tissues and could be useful for plant genetic engineering.