Knockdown of Sly-miR164a Enhanced Plant Salt Tolerance and Improved Preharvest and Postharvest Fruit Nutrition of Tomato.

Salinity stress is a serious limitation to tomato growth and development. The aim of this study was to investigate the effects of Sly-miR164a on tomato growth and fruit nutritional quality under salt stress. The results showed that the root length, fresh weight, plant height, stem diameter and ABA content of miR164a#STTM (knockdown of Sly-miR164a) lines were higher than those of WT and miR164a#OE (overexpression of Sly-miR164a) lines under salt stress. Compared with WT, miR164a#STTM tomato lines exhibited lower ROS accumulation under salt stress. In addition, the fruits of miR164a#STTM tomato lines had higher soluble solids, lycopene, ascorbic acid (ASA) and carotenoid content compared with WT. The study indicated that tomato plants were more sensitive to salt when Sly-miR164a was overexpressed, while knockdown of Sly-miR164a enhanced plant salt tolerance and improved fruit nutritional value.

Silencing of Sly-miR171d increased the expression of GRAS24 and enhanced postharvest chilling tolerance of tomato fruit.

The role of Sly-miR171d on tomato fruit chilling injury (CI) was investigated. The results showed that silencing the endogenous Sly-miR171d effectively delayed the increase of CI and electrolyte leakage (EL) in tomato fruit, and maintained fruit firmness and quality. After low temperature storage, the expression of target gene GRAS24 increased in STTM-miR171d tomato fruit, the level of GA3 anabolism and the expression of CBF1, an important regulator of cold resistance, both increased in STTM-miR171d tomato fruit, indicated that silencing the Sly-miR171d can improve the resistance ability of postharvest tomato fruit to chilling tolerance.

Chilling injury of tomato fruit was alleviated under low-temperature storage by silencing Sly-miR171e with short tandem target mimic technology.

In this study, the role of Sly-miR171e on post-harvest cold tolerance of tomato fruit was researched. The results showed that overexpression of Sly-miR171e (miR171e-OE) promoted postharvest chilling injury (CI) of tomato fruit at the mature red (MR) and mature green (MG) stage. Contrasted with the wild type (WT) and miR171e-OE fruit, the knockdown of Sly-miR171e (miR171e-STTM) showed a lower CI index, lower hydrogen peroxide (H2O2) content, and higher fruit firmness after harvest. In the fruit of miR171e-STTM, the expression level of GRAS24, CBF1, GA2ox1, and COR, and the GA3 content were ascended, while the expression levels of GA20ox1 and GA3ox1 were descended. The research demonstrated that CI in tomato fruit was alleviated at low temperature storage by silencing Sly-miR171e with short tandem target mimic (STTM) technology. Furthermore, it also provided helpful information for genetic modification of miR171e and control of CI in the postharvest fruit.

Melatonin maintained higher contents of unsaturated fatty acid and cell membrane structure integrity in banana peel and alleviated postharvest chilling injury.

Exogenous melatonin confers the chilling tolerance of banana fruit by promoting reactive oxygen species (ROS) scavenging system and inducing the unsaturated fatty acid synthesis. The results showed that melatonin treatment increased the contents of phospholipids, promoted the ROS scavenging enzyme, and restrained the activities of lipoxygenase (LOX), and thus reduced the lipid peroxidation of banana peel. In addition, melatonin treatment increased the flavonoids and proline contents, which was conducive to antioxidant capacity. Interestingly, the enhanced antioxidant capacity is conducive to the stability of unsaturated fatty acids and reduce the enzymatic browning reaction. Moreover, melatonin treatment induced the expression of omega-3/6 fatty acid desaturase and triggered the fatty acid metabolism activity, by which maintained higher contents of unsaturated fatty acid in banana peel. Moreover, melatonin treatment stimulated the accumulation of fatty acids in banana peel, and was involved in alleviating fruit chilling injury.

Dynamic Changes of DNA Methylation Induced by Heat Treatment Were Involved in Ethylene Signal Transmission and Delayed the Postharvest Ripening of Tomato Fruit.

Deoxyribonucleic acid (DNA) methylation plays an important role in fruit ripening and senescence. Here, the role of DNA methylation of the CpG island of SlACS10, LeCTR1, LeEIN3, LeERT10, and SlERF-A1 genes induced by heat treatment (37 °C) in postharvest ripening of tomato fruit was studied. After heat treatment, the firmness and vitamin C content showed higher levels, the loss of aldehydes in volatile components was delayed, and the activities of methylase and demethylase decreased in tomato fruit. Moreover, in heat-treated fruit, significant changes in DNA methylation of SlACS10, LeCTR1, LeEIN3, LeERT10, and SlERF-A1 were induced, the expression of LeERT10 and LeEIN3 was inhibited, the expression of SlERF-A1 was increased, by which ethylene signal transmission might be suppressed and the postharvest ripening of tomato fruit was delayed. The present study provided valuable information for understanding the essential role of DNA methylation in the postharvest ripening of tomato fruit.

Antifungal activity of 1-methylcyclopropene (1-MCP) against anthracnose (Colletotrichum gloeosporioides) in postharvest mango fruit and its possible mechanisms of action.

Anthracnose caused by Colletotrichum gloeosporioides is one of the most important postharvest diseases in mango fruit, often causing huge economic losses. In this study, the effect of 1-methylcyclopropene (1-MCP) against anthracnose in postharvest mango fruit and the mechanisms involved were investigated. 1-MCP induced reactive oxygen species (ROS) generation, damaged the mitochondria and destroyed the integrity of plasma membrane of spores of C. gloeosporioides, significantly suppressing spore germination and mycelial growth of C. gloeosporioides. 1-MCP also decreased the decay incidence and lesion expansion of mango fruit caused by C. gloeosporioides. For the first time this study demonstrated that 1-MCP suppressed anthracnose of postharvest mango fruit by directly inhibiting spore germination and mycelial growth of C. gloeosporioides, thus providing a promising strategy for disease control.

Deep sequencing identifies tissue-specific microRNAs and their target genes involving in the biosynthesis of tanshinones in Salvia miltiorrhiza.

Salvia miltiorrhiza is one of the most popular traditional medicinal herbs in Asian nations. Its dried root contains a number of tanshinones, protocatechuic aldehyde, salvianolic acid B and rosmarinic, and is used for the treatment of various diseases. The finding of microRNAs (miRNAs) and their target genes will help understand their biological role on the biosynthesis of tanshinones in S. miltiorrhiza. In the present study, a total of 452 known miRNAs corresponding to 589 precursor miRNAs (pre-miRNAs), and 40 novel miRNAs corresponding to 24 pre-miRNAs were identified in different tissues of S. miltiorrhiza by high-throughput sequencing, respectively. Among them, 62 miRNAs express only in root, 95 miRNAs express only in stem, 19 miRNAs express only in leaf, and 71 miRNAs express only in flower, respectively. By the degradome analysis, 69 targets potentially cleaved by 25 miRNAs were identified. Among them, acetyl-CoA C-acetyltransferase was cleaved by miR5072, and involved in the biosynthesis of tanshinones. This study provided valuable information for understanding the tissue-specific expression patterns of miRNAs in S. miltiorrhiza, and offered a foundation for future studies of the miRNA-mediated biosynthesis of tanshinones.

High-throughput sequencing and degradome analysis identify miRNAs and their targets involved in fruit senescence of Fragaria ananassa.

In non-climacteric fruits, the respiratory increase is absent and no phytohormone is appearing to be critical for their ripening process. They must remain on the parent plant to enable full ripening and be picked at or near the fully ripe stage to obtain the best eating quality. However, huge losses often occur for their quick post-harvest senescence. To understanding the complex mechanism of non-climacteric fruits post-harvest senescence, we constructed two small RNA libraries and one degradome from strawberry fruit stored at 20°C for 0 and 24 h. A total of 88 known and 1224 new candidate miRNAs, and 103 targets cleaved by 19 known miRNAs families and 55 new candidatemiRNAs were obtained. These targets were associated with development, metabolism, defense response, signaling transduction and transcriptional regulation. Among them, 14 targets, including NAC transcription factor, Auxin response factors (ARF) and Myb transcription factors, cleaved by 6 known miRNA families and 6 predicted candidates, were found to be involved in regulating fruit senescence. The present study provided valuable information for understanding the quick senescence of strawberry fruit, and offered a foundation for studying the miRNA-mediated senescence of non-climacteric fruits.

Seed size is determined by the combinations of the genes controlling different seed characteristics in rice.

Rice seed size is an important agronomic trait in determining the yield potential, and four seed size related genes (GS3, GW2, qSW5/GW5 and GIF1) have been cloned in rice so far. However, the relationship among these four genes is still unclear, which will impede the process of gene pyramiding breeding program to some extent. To shade light on the relationship of above four genes, gene expression analysis was performed with GS3-RNAi, GW2-RNAi lines and CSSL of qSW5 at the transcriptional level. The results clearly showed that qSW5 and GW2 positively regulate the expression of GS3. Meanwhile, qSW5 can be down-regulated by repression of GW2 transcription. Additionally, GIF1 expression was found to be positively regulated by qSW5 but negatively by GW2 and GS3. Moreover, the allelic effects of qSW5 and GS3 were detailedly characterized based on a natural population consisting of 180 rice cultivars. It was indicated that mutual interactions exist between the two genes, in which, qSW5 affecting seed length is masked by GS3 alleles, and GS3 affecting seed width is masked by qSW5 alleles. These findings provide more insights into the molecular mechanisms underlying seed size development in rice and are likely to be useful for improving rice grain yield.

Exogenous gamma-aminobutyric acid alleviates oxidative damage caused by aluminium and proton stresses on barley seedlings.

BACKGROUND: Proton (H(+)) and aluminium (Al(3+)) toxicities are major factors limiting crop production on acid soils, while gamma-aminobutyric acid (GABA) is a non-protein amino acid involved in various stress tolerances in plants. In this study, to determine whether exogenous GABA is functional in alleviating oxidative stress induced by H(+) and Al(3+) toxicities, the antioxidant defence response regulated by GABA was investigated in barley (Hordeum vulgare L.). RESULTS: After 24 h treatments of seedlings under H(+), Al(3+) and combined stresses with and without GABA, morphological and biochemical assays were conducted. It was observed that the inhibition of seedling root elongation caused by Al(3+) and H(+) toxicities was significantly mitigated by GABA. The amount of carbonylated proteins with molecular masses of 14.4-97 kDa was decreased. The activities of antioxidant enzymes were enhanced, the content of malondialdehyde was reduced and the accumulation of reactive oxygen species (ROS), as shown by staining roots with nitroblue tetrazolium, declined in GABA-treated seedlings. CONCLUSION: GABA can alleviate oxidative damage caused by H(+) and Al(3+) toxicities in barley seedlings by activating antioxidant defence responses and reducing the elevated levels of carbonylated proteins caused by ROS.

A 796 bp PsPR10 gene promoter fragment increased root-specific expression of the GUS reporter gene under the abiotic stresses and signal molecules in tobacco.

A 1681 bp PsPR10 promoter was isolated from Pinus strobus and a series of 5'-deletions were fused to the ß-glucuronidase (GUS) reporter gene and introduced into tobacco. GUS activity in P796 (-796 to +69) construct transgenic plant roots was similar with that of P1681 and higher than those of the P513 (-513 to +69) and P323 (-323 to +69) transgenic plants. Moreover, the abiotic stresses of NaCl, PEG 6000 and mannitol, and salicylic acid (SA), abscisic acid (ABA) and jasmonic acid (JA) induced higher GUS activity in the roots of P796 transgenic tobacco. This study provides a potential inducible root-specific promoter for transgenic plants.

Expression of five AtHsp90 genes in Saccharomyces cerevisiae reveals functional differences of AtHsp90s under abiotic stresses.

The genome of Arabidopsis thaliana contains seven Hsp90 family genes. Three organellar and two cytosolic AtHsp90 isoforms were characterized by functionally expressing them in a temperature-sensitive Hsp90 mutant and a conditional Hsp90-null mutant of Saccharomyces cerevisiae. The cytosolic AtHsp90-1 and AtHsp90-2 showed function similar to that of yeast in chaperoning roles; they could support the growth of yeast mutants at both permissive and non-permissive temperature. Neither the full-length nor mature forms of chloroplast-located AtHsp90-5, mitochondria-located AtHsp90-6 and endoplasmic reticulum (ER)-located AtHsp90-7 could complement the yeast Hsp90 proteins. The cytosolic AtHsp90s could stabilize the biomembrane of the temperature-sensitive Hsp90 mutant strains under stress conditions, while the organellar AtHsp90s could not protect the biomembrane of the temperature-sensitive Hsp90 mutant strains. Yeast two-hybrid results showed that either pre-protein or mature forms of organellar AtHsp90s could interact with cofactors cpHsp70, Hsp70, Hsp70t-2, Cyp40, p23 and a substrate protein of NOS, while cytosolic AtHsp90s could not interact with them. These results suggest that organellar and cytosolic AtHsp90s possibly work through different molecular mechanisms in forming chaperone complexes and performing their functional roles.

Functional characterization of AtHsp90.3 in Saccharomyces cerevisiae and Arabidopsis thaliana under heat stress.

The function of cytosolic AtHsp90.3 was characterized by complementing the Saccharomyces cerevisiae endogenous Hsp90 genes and overexpressing it in Arabidopsis thaliana. Though AtHsp90.3 supported the yeast growth under heat stress, in Arabidopsis, compared to the wild type, the transgenic plants overexpressing cytosolic AtHsp90.3 were more sensitive to heat stress with a lower germination rate and higher mortality but and more tolerant to high Ca(2+). Transcriptional expression of heat stress transcription factors, AtHsfA1d, AtHsfA7a and AtHsfB1, and two Hsps, AtHsp101 and AtHsp17, was delayed by constitutive overexpression of cytosolic AtHsp90.3 under heat stress. These results indicate that overexpressing AtHsp90.3 impaired plant tolerance to heat stress and proper homeostasis of Hsp90 was critical for cellular stress response and/or tolerance in plants.

Comparative proteomic analysis of differentially expressed proteins in shoots of Salicornia europaea under different salinity.

Soil salinity is a major abiotic stress that limits agriculture productivity worldwide. Salicornia europaea is a succulent annual euhalophyte and one of the most salt tolerant plant species. The elucidation of its salt tolerance mechanism is of significance for generating salt-tolerant crops. In this study, we provided high resolution of proteome reference maps of S. europaea shoot and obtained evidence on the salt tolerance mechanism by analyzing the proteomic responses of this plant to high salinity. Our results demonstrated significant variations existed in 196 out of 1880 protein spots detected on CBB stained 2-DE gels. Of these, 111 proteins were identified by mass spectrometry. Among them, the majority was energy production and conversion related proteins, followed by photosynthesis and carbohydrate metabolism associated enzymes. Analysis of protein expression patters revealed that energy production and ion homeostasis associated proteins played important roles for this plant salt tolerance ability. Hierarchical clustering results revealed many proteins were involved in S. europaea salt tolerance mechanism as a dynamic network. Finally, based on our proteomic results, we brought forward a possible schematic representation of mechanism associated with the systematic salt tolerance phenotype in S. europaea.

Overexpression of AtHsp90.2, AtHsp90.5 and AtHsp90.7 in Arabidopsis thaliana enhances plant sensitivity to salt and drought stresses.

Three AtHsp90 isoforms, cytosolic AtHsp90.2, chloroplast-located AtHsp90.5, and endoplasmic reticulum (ER)-located AtHsp90.7, were characterized by constitutive overexpressing their genes in Arabidopsis thaliana. Both types of the transgenic plants overexpressing cytosolic and organellar AtHsp90s showed reduced tolerance to salt and drought stresses with lower germination rates and fresh weights, but improved tolerance to high concentration of Ca(2+) comparing with the wild type plants. Transcriptional analysis of ABA-responsive genes, RD29A, RD22 and KIN2 under salt and drought stresses, indicated that the induction expression of these genes was delayed by constitutive overexpression of cytosolic AtHsp90.2, but was hardly affected by that of organellar AtHsp90.5 and AtHsp90.7. These results implied that Arabidopsis different cellular compartments-located Hsp90s in Arabidopsis might be involved in abiotic stresses by different functional mechanisms, probably through ABA-dependent or Ca(2+) pathways, and proper homeostasis of Hsp90 was critical for cellular stress response and/or tolerance in plants.