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Key message: We report that GhWRKY21, a WRKY transcription factor, plays essential roles in regulating the intensity of the drought-induced ABA signalling pathway by facilitating the expression of GhHAB in cotton (Gossypium hirsutum). Abscisic acid (ABA) is one of the most important plant hormones in response to abiotic stress. However, activation of the ABA signalling pathway often leads to growth inhibition. The mechanisms that regulate the intensity of ABA signals are poorly understood. Here, we isolated and analysed the cotton group IId WRKY transcription factor (TF) gene GhWRKY21. Functional analysis indicated that GhWRKY21 plays a negative role in the drought response of cotton. Silencing of GhWRKY21 in cotton dramatically increased drought tolerance, whereas ectopic GhWRKY21 overexpression in Nicotiana benthamiana decreased drought tolerance. Furthermore, the GhWRKY21-mediated drought tolerance was ABA dependent. To clarify the mechanism underlying the GhWRKY21-mediated regulation of drought tolerance, 17 clade-A-type type 2C protein phosphatase (PP2C) genes, which are negative regulators of ABA signalling, were identified in cotton. Notably, GhWRKY21 interacted specifically with the W-box element within the promoter of GhHAB and regulated its expression. Silencing of GhHAB in cotton yielded a phenotype similar to that of GhWRKY21-silenced cotton. These results suggest that GhWRKY21 regulates the intensity of ABA signals by facilitating the expression of GhHAB. In summary, these findings dramatically improve our understanding of the function of WRKY TFs and provide insights into the mechanism of ABA-mediated drought tolerance. [ABSTRACT FROM AUTHOR]

Bees are vitally important in natural and agricultural ecosystems, providing key pollination services to wild plants and crops. Increasing reports of regional declines of bee populations have attracted intense attention worldwide. Challenges to bee health are multifactorial and include poor nutrition, heat stress, agrochemicals, and pathogens. The impact of heat stress is a relatively minor factor in current bee declines compared with agrochemicals and pathogens. However, heat stress has adverse impacts on foraging activity, pollination services, task-related physiology, immunocompetence, reproductive capacity, growth, and development of bees, and these adverse impacts are variable in different bee species. Heat stress–related damage to bees receives extra attention when it is accompanied by climate change. Heat-tolerance mechanisms are key enablers for bee survival under high-temperature stress conditions, and we now understand that both behavior and molecular regulation strongly impact the ability of bees to reduce damage from heat stress. In this review, we summarize and synthesize previous findings about the detrimental effects of heat stress to bees and discuss the strategies bees use to cope with heat stress. Bee species mentioned here are mainly honeybees, bumblebees, and stingless bees, with a focus on the honeybee Apis mellifera. [ABSTRACT FROM AUTHOR]

As predominant pollinators, honeybees are important for crop production and terrestrial ecosystems. Recently, various environmental stresses have led to large declines in honeybee populations in many regions. The ability of honeybees to respond to these stresses is critical for their survival. However, the details of the stress defense mechanisms of honeybees have remained elusive. Here, we found that the Nuclear Factor Y (NF-Y) family (containing NF-YA, NF-YB, and NF-YC) is a novel stress mediator family that regulates honeybee environmental stress resistance. NF-YA localized in the nucleus, NF-YB accumulated in the cytoplasm, and NF-YC presented in both the nucleus and cytoplasm. NF-YC interacted with NF-YA and NF-YB in vitro and in vivo, and the nuclear import of NF-YB relied on its interaction with NF-YC. We further found that the expression of NF-Y was induced under multiple stress conditions. In addition, NF-Y regulated many stress responses and antioxidant genes at the transcriptome-wide level, and knockdown of NF-Y repressed the expression of stress-inducible genes, particularly LOC108003540 and LOC107994062, under adverse circumstances. Silencing NF-Y lowered honeybee stress resistance by reducing total antioxidant capacity and enhancing oxidative impairment. Collectively, these results indicate that NF-Y plays important roles in stress responses. Our study sheds light on the underlying defense mechanisms of honeybees under environmental stress. [ABSTRACT FROM AUTHOR]

Vitellogenin (Vg) is a yolk precursor protein in most oviparous females. However, Vg has not been studied in the Apis cerana cerana. In this work, the Vg gene of the A. cerana cerana has been cloned and sequenced. The gene codes for a protein consisting of 1770 amino acids in seven exons with a predicted molecular mass and isoelectric point of 200 kDa and 6.46, respectively. Additionally, an 807-bp 5′-flanking region was isolated, and potential transcription factor binding sites associated with development and stress response were identified. Quantitative real-time PCR analysis showed that AccVg is highly expressed in pupae during different developmental stages. In addition, the expression of AccVg could be induced by cold (4 and 16 °C), CdCl, and pesticide treatment. Real-time quantitative PCR (RT-PCR) and western blot analyses indicated that AccVg transcription was induced by various abiotic stresses. Western blot was used to measure the expression levels of AccVg protein. Taken together, these results suggest that AccVg most likely plays essential roles in antioxidant defence and that it may be of critical importance to honey bee survival. [ABSTRACT FROM AUTHOR]

Apis cerana cerana triosephosphate isomerase ( AccTPI), a key regulatory enzyme in glycolysis and gluconeogenesis pathway, catalyzes the interconversion of glyceraldehyde 3-phosphate to dihydroxyacetone phosphate. However, its role in A. cerana cerana has not been completely clarified. In this study, a TPI gene was cloned from A. cerana cerana and named AccTPI. The open reading frame (ORF) of AccTPI is 744 bp, which encodes 247 amino acids with a predicted molecular weight and an isoelectric point of 26.8 kDa and 8.42, respectively. The expression of AccTPI was up-regulated by some abiotic stresses, while it was down-regulated by HgCl, ultraviolet light (UV), and vitamin C (VC) treatment. Finally, a disc diffusion assay revealed that recombinant AccTPI proteins can protect cells from oxidative stresses. Taken together, all of the results indicate that AccTPI may play an important role in the metabolism, in antioxidant defense, and in many biological functions in the growth and development of A. cerana cerana. [ABSTRACT FROM AUTHOR]

Apoptosis-inducing factors (AIFs) play vital roles in bioenergetic and redox metabolism. Compared with studies focused on the apoptogenic function of AIF, few studies have reported on its oxidoreductase function, especially in insects. In this study, we identified a novel AIF gene AccAIF3 that was isolated from Apis cerana cerana, and investigated its expression and structural features. Quantitative real-time PCR (qRT-PCR) revealed the highest mRNA level of AccAIF3 in adult bees at 15 days. qRT-PCR results revealed that AccAIF3 transcripts were induced by cold, CdCl, HgCl, UV, pyriproxyfen and cyhalothrin, but were downregulated by ecdysone. We also compared the expression of AIF3 from Apis cerana cerana and Apis mellifera ligustica Spinola to Ascosphaera apis, and found that AIF3s may respond to Ascosphaera apis in a species-specific manner. These data suggested that AccAIF3 may play vital roles in the response to abiotic and biotic stresses and contribute to the adaptability of honeybees to adversities. [ABSTRACT FROM AUTHOR]

Key message: Our results indicate that overexpression of the GhWRKY39 - 1 gene enhances resistance to pathogen infection and tolerance to high salt and oxidative stress in transgenic Nicotiana benthamiana. Abstract: WRKY transcription factor genes play significant roles in the response to biotic and abiotic stresses. Cotton ( Gossypium hirsutum) is an important fiber and oil crop worldwide. We isolated and characterized GhWRKY39- 1, which is a group IId WRKY gene that is present as a single copy in the cotton genome. Quantitative PCR analyses indicated that GhWRKY39- 1 was induced by pathogen infection, defense-related signaling molecules, and abiotic stresses, such as NaCl and methyl viologen. An analysis of the subcellular localization of the GhWRKY39-1 protein indicated that it localized to the nucleus. Furthermore, constitutive overexpression of GhWRKY39- 1 in Nicotiana benthamiana conferred a greater resistance to infection by both the bacterial pathogen Ralstonia solanacearum and the fungal pathogen Rhizoctonia solani. The transgenic plants also exhibited elevated mRNA levels of several pathogen-related ( PR) genes, including PR1c, PR2 and PR4. Moreover, transgenic plants displayed an enhanced tolerance to salt and oxidative stress and elevated expression of several oxidation-related genes, including APX, CAT, GST and SOD. Overall, these results indicate that GhWRKY39- 1 functions as a positive regulator of plant defense against pathogen infection and responses to salt stress and reactive oxygen species. [ABSTRACT FROM AUTHOR]

RNA-dependent RNA polymerases (RDRs) play key roles in RNA silencing, developmental regulation, antiviral defense and the maintenance of genome integrity. In this work, we isolated an RDR gene ( NtRDR6) from Nicotiana tabacum using reverse-transcription PCR (RT-PCR) and rapid amplification of cDNA ends PCR (RACE-PCR) technology. The full-length cDNA of NtRDR6 encodes a protein of 1,197 amino acids, which contains all of the five highly conserved regions in plant RDRs and the DxDGD signature motif. An amino acid sequence alignment revealed that NtRDR6 shares a high level of similarity with other known RDRs. The genomic sequence of NtRDR6 contains two introns and a group of putative cis-acting elements in the 5′-flanking region. Expression analysis performed using 6-week-old seedlings revealed that the transcript level of NtRDR6 is up-regulated by diverse signalling molecules, including jasmonate (JA), abscisic acid (ABA), gibberellin A (GA) and salicylic acid (SA). The expression of NtRDR6 is also elevated markedly by Cucumber mosaic virus (CMV) and Potato virus X (PVX) but not by Potato virus Y (PVY) and Tobacco mosaic virus (TMV). Notably, abiotic stresses have alternative effects on the transcription of NtRDR6, with heat (37°C) significantly increasing the expression of NtRDR6, whereas low temperature (15°C) does not affect expression. These results indicated that NtRDR6 may be involved in several signal transduction pathways to defend against plant virus attack and in the response to abiotic stresses. [ABSTRACT FROM AUTHOR]

The cDNA of AtPLDα (Arabidopsis thaliana Phospholipase Dα) gene was introduced into P. tomentosa (Populus tomentosa) under the control of the Cauliflower mosaic virus 35S promoter. Southern and Northern blot analyses suggested that the AtPLDα gene has been transferred into the P. tomentosa genome. No obvious morphological or developmental difference was observed between the transgenic and wild-type (WT) plants. Drought and salt tolerance and gene expression of seedlings of several transgenic lines and WT plants (control) were studied. The results showed that the rhizogenesis rate and the average root-length of transgenic lines were significantly higher than WT plants after mannitol and NaCI treatment under the same growth conditions. Northern blot analysis indicated that the higher the PLDα expression in the transgenic plants, the more tolerant the transgenic plants are to drought and salt treatment. Meanwhile, another group of these transgenic lines and WT plants (control) were treated with PEG6000 and NaCI separately. The contents of chlorophylls and the activities of some antioxidant enzymes (superoxide dismutase, guaiacol peroxidase and catalase) as well as malondialdehyde and relative electrical conductivity were analyzed. Altogether, our results demonstrated that overexpression of the PLDα gene can enhance the drought and salt tolerance in transgenic P. tomentosa plants. [ABSTRACT FROM AUTHOR]

Reports that the inclusion of MARs in transgene cassettes enhances their expression and reduces position-effect variations in the transgenic host. Statement that four new MARs (TM2, TM3, AM1 and AM2) were isolated from tobacco and Arabidopsis by PCR method; Reference to nuclei isolated from suspensioncultured cells of rice used to prepare nuclear matrix; Report that with a characterized MAR (TM1) as a positive control, the Matrix-MAR interactions were tested by an in vitro binding assay to identify the DNA sequences as MARs and their binding strength to nuclear matrix in vitro was compared; Results showing that TM2 and TM3 had stronger binding strength than TM1; Procedure used to determine the functions of the four new MARs in vivo; Modification of binary vectors pBI121 carrying a u/dA GUS reporter gene with direct repeat MARs inserted on both sides of the reporter gene cassette; Transferance into tobaccos via Agrobacterium-mediated transformation procedure. Statement that quantitative GUS assays of the transgenie tobaccos showed that when flanking a GUS reporter gene TM1, TM2, TM3 and AM1 increased u/dA GUS gene expression level approximately 1.5-fold, 5-fold, 1.35-fold, 1.3-fold respectively and AM2 has no effect on gene expression. Discovery that TM2 was found to be a strong MAR that could effectively increase gene expression level and could be used as an effective enhancing element to construct high efficient expression vectors; Discussion of the relations among the sequence features, binding strength in vitro and function in vivo of the five MARs that were analyzed, and the potential significance of TM2 in plant genetic engineering.