Genetic transformation of cotton with a harpin-encoding gene hpaXoo confers an enhanced defense response against different pathogens through a priming mechanism

<p>Abstract</p> <p>Background</p> <p>The soil-borne fungal pathogen <it>Verticillium dahliae </it>Kleb causes <it>Verticillium </it>wilt in a wide range of crops including cotton (<it>Gossypium hirsutum</it>). To date, most upland cotton varieties are susceptible to <it>V. dahliae </it>and the breeding for cotton varieties with the resistance to <it>Verticillium </it>wilt has not been successful.</p> <p>Results</p> <p>Hpa1_Xoois a harpin protein from <it>Xanthomonas oryzae </it>pv. <it>oryzae </it>which induces the hypersensitive cell death in plants. When <it>hpa1</it>_{<it>Xoo </it>}was transformed into the susceptible cotton line Z35 through <it>Agrobacterium</it>-mediated transformation, the transgenic cotton line (T-34) with an improved resistance to <it>Verticillium dahliae </it>was obtained. Cells of the transgenic T-34, when mixed with the conidia suspension of <it>V. dahliae</it>, had a higher tolerance to <it>V. dahliae </it>compared to cells of untransformed Z35. Cells of T-34 were more viable 12 h after mixing with <it>V. dahliae </it>conidia suspension. Immunocytological analysis showed that Hpa1_Xoo, expressed in T-34, accumulated as clustered particles along the cell walls of T-34. In response to the infection caused by <it>V. dahliae</it>, the microscopic cell death and the generation of reactive oxygen intermediates were observed in leaves of T-34 and these responses were absent in leaves of Z35 inoculated with <it>V. dahliae</it>. Quantitative RT-PCR analysis indicated that five defense-related genes, <it>ghAOX1, hin1, npr1, ghdhg-OMT</it>, and <it>hsr203J</it>, were up-regulated in T-34 inoculated with <it>V. dahliae</it>. The up-regulations of these defense-relate genes were not observed or in a less extent in leaves of Z-35 after the inoculation.</p> <p>Conclusions</p> <p>Hpa1_Xooaccumulates along the cell walls of the transgenic T-34, where it triggers the generation of H₂O₂as an endogenous elicitor. T-34 is thus in a primed state, ready to protect the host from the pathogen. The results of this study suggest that the transformation of cotton with <it>hpa1</it>_{<it>Xoo </it>}could be an effective approach for the development of cotton varieties with the improved resistance against soil-borne pathogens.</p

Anti-inflammatory Effects of α7-nicotinic ACh Receptors are Exerted Through Interactions with Adenylyl Cyclase-6

Background and purpose Alpha 7 nicotinic acetylcholine receptors (CHRNA7) suppress inflammation through diverse pathways in immune cells, so is potentially involved in a number of inflammatory diseases. However, the detailed mechanisms underlying CHRNA7’s anti‐inflammatory effects remain elusive. Experimental approach The anti‐inflammatory effects of CHRNA7 agonists in both murine macrophages (RAW 264.7) and bone marrow‐derived macrophages (BMDM) stimulated with LPS were examined. The role of adenylyl cyclase 6 (AC6) in Toll‐like Receptor 4 (TLR4) degradation was explored via overexpression and knockdown. A mouse model of chronic obstructive pulmonary disease was used to confirm key findings. Results Anti‐inflammatory effects of CHRNA7 were largely dependent on AC6 activation, as knockdown of AC6 considerably abnegated the effects of CHRNA7 agonists while AC6 overexpression promoted them. We found that CHRNA7 and AC6 are co‐localized in lipid rafts of macrophages and directly interact. Activation of AC6 led to the promotion of TLR4 degradation. Administration of CHRNA7 agonist PNU‐282987 attenuated pathological and inflammatory end points in a mouse model of chronic obstructive pulmonary disease (COPD). Conclusion and implications CHRNA7 inhibits inflammation through activating AC6 and promoting degradation of TLR4. The use of CHRNA7 agonists may represent a novel therapeutic approach for treating COPD and likely other inflammatory diseases

Modulation of cardiac resident macrophages immunometabolism upon high-fat-diet feeding in mice

BackgroundA high-fat diet (HFD) contributes to various metabolic disorders and obesity, which are major contributors to cardiovascular disease. As an essential regulator for heart homeostasis, cardiac resident macrophages may go awry and contribute to cardiac pathophysiology upon HFD. Thus, to better understand how HFD induced cardiac dysfunction, this study intends to explore the transcriptional and functional changes in cardiac resident macrophages of HFD mice.MethodsC57BL/6J female mice that were 6 weeks old were fed with HFD or normal chow diet (NCD) for 16 weeks. After an evaluation of cardiac functions by echocardiography, mouse hearts were harvested and cardiac resident CCR2- macrophages were sorted, followed by Smart sequencing. Bioinformatics analysis including GO, KEGG, and GSEA analyses were employed to elucidate transcriptional and functional changes.ResultsHyperlipidemia and obesity were observed easily upon HFD. The mouse hearts also displayed more severe fibrosis and diastolic dysfunction in HFD mice. Smart sequencing and functional analysis revealed metabolic dysfunctions, especially lipid-related genes and pathways. Besides this, antigen-presentation-related gene such as Ctsf and inflammation, particularly for NF-κB signaling and complement cascades, underwent drastic changes in cardiac resident macrophages. GO cellular compartment analysis was also performed and showed specific organelle enrichment trends of the involved genes.ConclusionDysregulated metabolism intertwines with inflammation in cardiac resident macrophages upon HFD feeding in mice, and further research on crosstalk among organelles could shed more light on potential mechanisms

MiR-34a Enhances Chondrocyte Apoptosis, Senescence and Facilitates Development of Osteoarthritis by Targeting DLL1 and Regulating PI3K/AKT Pathway

Background/Aims: Osteoarthritis (OA) is the prevalent degenerative disease caused by various factors. MicroRNAs are important regulators in the inflammation and immune response. The aim of this study was to investigate the effect of microRNA-34a (MiR-34a) on the death of chondrocytes, senescence, as well as its role in OA progression. Methods: A series of experiments involving CCK-8, flow cytometry, β-galactosidase staining and wound healing assays were conducted to determine the cellular capabilities of proliferation, cell apoptosis, senescence and the ability of cells to recover from injury, respectively. Binding sites between miR-34a and delta-like protein 1 (DLL1) were identified using a luciferase reporter system, whereas mRNA and protein expression of target genes was determined by RT-PCR and immunoblot, respectively. OA model was generated via surgery. Results: We found that miR-34a expression was increased in the cartilage of OA patients. In rat chondrocytes and chondrosarcoma cells, miR-34a transfections noticeably inhibited the expression of DLL1, triggered cell death and senescence, suppressed proliferation, and prevented scratch assay wound closure. However, transfection of a miR-34a inhibitor displayed adverse effects. Additionally, secretion and expression of factors associated with cartilage degeneration were altered via miR-34a. Moreover, miR-34a directly inhibits DLL1 mRNA. Furthermore, concentrations of DLL1, total PI3K, and p-AKT declined in chondrocytes that overexpress miR-34a. DLL1 overexpression elevated PI3K and p-AKT levels, and eliminated cell death triggered by a miR-34a mimic. In vivo, miR-34a remarkably inhibited miR-34a up-regulation, while enhanced the level of DLL1 expression. In the knee joints of surgery-induced OA rats, articular chondrocyte death and loss of cartilage were attenuated via miR-34a antagomir injection. Conclusions: These findings indicate that miR-34a contributes to chondrocyte death, causing OA progression through DLL1 and modulation of the PI3K/AKT pathway

Transcriptome analysis of Hpa1Xoo transformed cotton revealed constitutive expression of genes in multiple signalling pathways related to disease resistance

The transcriptome profile in leaves and roots of the transgenic cotton line T-34 expressing hpa1Xoo from Xanthomonas oryzae pv. oryzae was analysed using a customized 12k cotton cDNA microarray. A total of 530 cDNA transcripts involved in 34 pathways were differentially expressed in the transgenic line T-34, in which 123 differentially expressed genes were related to the cotton defence responses including the hypersensitive reaction, defence responses associated with the recognition of pathogen-derived elicitors, and defence signalling pathways mediated by salicylic acid, jasmonic acid, ethylene, auxin, abscicic acid, and Ca2+. Furthermore, transcripts encoding various leucine-rich protein kinases and mitogen-activated protein kinases were up-regulated in the transgenic line T-34 and expression of transcripts related to the energy producing and consuming pathway was also increased, which suggested that the enhanced metabolism related to the host defence response in the transgenic line T-34 imposed an increased energy demand on the transgenic plant

Characterization of Root Morphology and Anatomical Structure of Spring Maize under Varying N Application Rates and Their Effects on Yield

Root morphology is an important factor determining nitrogen (N) uptake by plants, which might be affected by the extent of N application. The processes associated with root morphogenesis of spring maize in response to N application rates remain poorly understood. In this study, both field and pot experiments were conducted to explore the effect of zero-N (N0), optimized-N (N180), and high-N (N360) on root morphology, anatomical structure, and N accumulation in spring maize. N application rates affected root length and surface area, and its endogenous hormone contents. The largest difference in total root length and surface area among the three N rates was found at the silking stage: the total root length and surface increased by 51.36% and 42.58% under N180 and by 7.8% and 30.14% under N360, respectively, compared with N0, and the root/shoot ratio and root bleeding sap significantly increased under N180 and N360 compared with N0. The auxin and jasmonic acid levels of roots under N180 and N360 were higher than N0. N application rates also affected root microstructure and ultrastructure. Compared with N0, the proportions of root aerating tissue under N180 and N360 were decreased by 32.42% and 11.92% at silking. The root tip cell structure was damaged under N0, and intact under N180 and N360. Moreover, the 15N allocation proportions to root and grain under N180 and N360 were increased compared to N0. Grain yields under N180 and N360 increased by 20.44% and 16.6% compared with N0, respectively. It can be concluded that optimized-N application decreased root aerated tissue and thus improved root length and root surface area through regulating auxin and jasmonic acid levels and affected N uptake and grain yield of N-efficient spring maize variety