Molecular cloning of a novel chimeric HMW glutenin subunit gene <Emphasis Type="Italic">1Dx5′</Emphasis> from a common wheat line W958

A novel chimeric high-molecular-weight (HMW) glutenin subunit gene from a new common wheat line W958 (2n = 6x = 42) was isolated and characterized. SDS–PAGE analysis revealed that this glutenin subunit has similar electrophoretic mobility to 1Dx5, so it was designated 1Dx5′. Genomic DNA from W958 was amplified and a 2,505-bp fragment was obtained. The 1Dx5′ subunit showed a chimeric primary structure of 1Dx5 and 1Dx2, with the 1Dx5 sequence in the 5′ and middle repetitive regions and the 1Dx2 sequence in the repetitive domain and 3′ region. MALDI-TOF-MS analysis demonstrated that 1Dx5′ had a molecular weight of 86815.1 Da, close to that of an x-type glutenin subunit. Secondary structure analysis showed that this subunit had six helixes and one strand, including four helixes in the repetitive domain which could enhance the dough properties. Additionally, the promoter of 1Dx5′ was obtained and showed the same sequence as 1Dx5 or 1Dx2 except for a few base conversions. The promoter analysis indicated that the cis-acting regulatory elements of 1Dx5′ were the same as those of 1Dx5 and/or 1Dx2. Previously, we have demonstrated that this novel glutenin subunit is associated with good bread-making quality and comprises a very large proportion of the F2 segregation population. Consequently, we suggest that the amino acid residue composition and the secondary structure of the subunit may contribute to the bread-making quality. In summary, the novel 1Dx5′ gene could have greater potential in wheat quality improvement.     

The cis-expression of the coat protein of turnip mosaic virus is essential for viral intercellular movement in plants

To establish infection, plant viruses are evolutionarily empowered with the ability to spread intercellularly. Potyviruses represent the largest group of known plant-infecting RNA viruses, including many agriculturally important viruses. To better understand intercellular movement of potyviruses, we used turnip mosaic virus (TuMV) as a model and constructed a double-fluorescent (green and mCherry) protein-tagged TuMV infectious clone, which allows distinct observation of primary and secondary infected cells. We conducted a series of deletion and mutation analyses to characterize the role of TuMV coat protein (CP) in viral intercellular movement. TuMV CP has 288 amino acids and is composed of three domains: the N-terminus (amino acids 1–97), the core (amino acids 98–245), and the C-terminus (amino acids 246–288). We found that deletion of CP or its segments amino acids 51–199, amino acids 200–283, or amino acids 265–274 abolished the ability of TuMV to spread intercellularly but did not affect virus replication. Interestingly, deletion of amino acids 6–50 in the N-terminus domain resulted in the formation of aberrant virions but did not significantly compromise TuMV cell-to-cell and systemic movement. We identified the charged residues R178 and D222 within the core domain that are essential for virion formation and TuMV local and systemic transport in plants. Moreover, we found that trans-expression of the wild-type CP either by TuMV or through genetic transformation-based stable expression could not rescue the movement defect of CP mutants. Taken together these results suggest that TuMV CP is not essential for viral genome replication but is indispensable for viral intercellular transport where only the cis-expressed CP is functional.     

救护白鹤与野生白鹤迁徙路线和活动规律差异分析

受伤和体弱白鹤的救护及放归是白鹤保护的重要方式之一。2014—2016年间,先后对8只救护白鹤和2只野生白鹤进行环志并安装卫星跟踪器,根据所获得的卫星跟踪数据来分析比较救护白鹤与野生白鹤在秋季迁徙路线和在重要中途停歇地的最大日活动距离、活动区分布和面积、生境类型等方面的异同。结果显示,救护白鹤放归当年与放归次年以及救护白鹤放归当年与野生白鹤的秋季迁徙路线和活动规律之间存在明显差异,但救护白鹤放归次年与野生白鹤之间差异不明显。(1)放归当年,4只救护白鹤在农田和水塘停歇1个月左右,停歇时长明显高于野生白鹤和放归次年的救护白鹤。(2)救护白鹤放归当年在跨海方式、登陆地点、停歇地点以及登陆后的迁徙方向上表现出多样性和不确定性,其中部分个体在迁徙过程中有停顿徘徊、迷失迁徙方向的现象,且跨越渤海和大别山的距离大于放归次年的救护白鹤和野生白鹤。(3)救护白鹤放归后4天内每日最大活动距离小,而后高低分化明显,无明显规律,而救护白鹤放归次年的每日最大活动距离规律变化与野生白鹤较一致。(4)与救护白鹤放归次年以及野生白鹤的活动规律相比,救护白鹤放归当年的栖息地位点变化频繁,具体表现为生境类型选择的多样化,且无明显规律,同时活动区面积较小。探索救护白鹤放归野外后与野生白鹤迁徙路线和活动规律的差异,对救护白鹤的科学放归具有指导意义。     

Effect of Host Plants on Honeydew Production of an Invasive Mealybug, Phenacoccus solenopsis (Hemiptera: Pseudococcidae)

Honeydew production plays a key role in mutualism between the mealybugs and ants. However, no studies have focused on the amount and circadian rules of honeydew excreted by Phenacoccus solenopsis Tinsley, a new invasive species which has conditional mutualism with Solenopsis invicta Buren in China. To address this problem, we measured the weight and estimated honeydew production in all stages of development of the invasive mealybug, P. solenopsis, as well as its honeydew production on tomato (Solanum lycopersicun), Hibiscus rosa-sinensis, and cotton (Gossypium sp.) for 24 h. The honeydew excreted by each instar of the mealybug in H. rosa-sinensis was measured for 2 weeks. Our results revealed that the weight of mealybugs significantly varied at different development stages. Host plants had no significant effect on the weight of nymphs, although the weight of a single adult reared on S. lycopersicun was significantly heavier than those reared on H. rosa-sinensis and G. sp. The amount of honeydew excreted by the 1st instar nymphs in S. lycopersicum was significantly greater than that on H. rosa-sinensis and G. sp. Each instar mealybug produced more honeydew when fed with S. lycopersicum compared with H. rosa-sinensis and G. sp. The amount of honeydew excreted by mealybugs when provisioned with H. rosa-sinensis was no different from mealybugs provisioned with G. spp. while in the same instar. The amount of honeydew excreted by the 1st and 2nd instar nymphs was not significantly different on the same host plant. However, there was a significant difference between the 3rd instar nymph and the adult. The amount of honeydew excreted by a single adult when provisioned with H. rosa-sinensis decreased from 3085.3 μg to 572.0 μg in 2 weeks. The 2nd instar nymph, 3rd instar nymph, and adult excreted honeydew more frequently during the day than at night, while the frequency of honeydew excretion of the 1st instar nymph had no significant difference between daytime and night.     

Expression and role of Toll-like receptors on human umbilical cord mesenchymal stromal cells

Background aimsToll-like receptors (TLRs) play an important role in innate and adaptive immunity by recognizing pathogen-associated molecular patterns (PAMPs).MethodsIn the present study, we investigated the expression and role of TLRs on human umbilical cord mesenchymal stromal cells (UC-MSCs). The proliferation, differentiation and immunoregulatory activity of UC-MSCs primed with or without TLR ligands were determined.ResultsAt the RNA level, the expression of TLR2, 4, 6 and 9 was relatively higher than that of other TLRs. However, TLR3 and TLR4 expression were relatively higher at the protein level. UC-MSCs expressed functional TLRs by nuclear factor-κB activation and cytokine expression assay. Poly-inosinic acid:cytidylic acid [Poly(I:C)] stimulation inhibited the proliferation of UC-MSCs, but the ligand of other TLRs had no significant effect. Poly(I:C) stimulation enhanced the adipogenic differentiation capability of UC-MSCs, but lipopolysaccharide inhibited the adipogenic differentiation. Poly(I:C) and CpG-oligonucleotide promoted the immunosuppressive potentiality of UC-MSCs, accompanied with the phosphorylation of interferon regulatory factor 3 (IRF3) and increased expression of indoleamine 2,3-dioxygenase and interferon β, whereas activation of other TLR ligands (synthetic analog fibroblast-stimulating lipopeptide-1 and lipopolysaccharide) failed to affect the immunoregulatory activity of UC-MSCs.ConclusionsTaken together, our data demonstrated that TLR activation influenced the function of UC-MSCs, which might have important implications in future efforts to explore the clinical potentials of UC-MSCs.     

Effects of Different Sources and Levels of Zinc on H2O2-Induced Apoptosis in IEC-6 Cells

Zinc has been shown to be an inhibitor of apoptosis for many years. The present study was designed to investigate effects of three zinc chemical forms on H₂O₂-induced cell apoptosis in IEC-6 cells via analysis of cell vitality, LDH activity, apoptosis percentage, caspase-3 activity, and Bcl-2, Bax, and caspase-3, -8, and -9 gene expression. Cells were divided into H₂O₂ and zinc sources+H₂O₂ groups, and there are three different zinc sources [zinc oxide nanoparticle (nano-ZnO), zinc oxide (ZnO), and zinc sulfate (ZnSO₄)] and three concentrations (normal = 25 μM, medium = 50 μM, and high = 100 μM) used in this article. In the present study, we found the striking cytotoxicity of H₂O₂ higher than 200 μM on cell vitality, LDH activity, and apoptosis percentage in the cells using five different concentrations (50, 100, 200, 400, and 800 μM) of H₂O₂ for 4 h. Moreover, we observed that cell vitality was increased, LDH activity and apoptotic percentage were decreased, and gene expression level of Bax and caspase-3 and -9 was markedly reduced, while gene expression level of Bcl-2 and ratio of Bcl-2/Bax were increased in normal concentration groups of nano-ZnO and ZnSO₄ compared with H₂O₂ group, but no significant difference was observed in caspase-8 gene expression. Furthermore, medium or, more intensely, high concentrations of nano-ZnO and ZnSO₄ enhanced H₂O₂-induced cell apoptosis. Compared with nano-ZnO and ZnSO₄, ZnO showed weakest protective effect on H₂O₂-induced apoptosis at normal concentration and was less toxic to cells at high level. Taken together, we proposed that preventive and protective effects of zinc on H₂O₂-induced cell apoptosis varied in IEC-6 cells with its chemical forms and concentrations, and maybe for the first time, we suggested that nano-ZnO have a protective effect on H₂O₂-induced cell apoptosis in IEC-6 cells.     

药源植物盾叶薯蓣甾体皂苷及皂苷元的研究进展

盾叶薯蓣是重要的甾体激素类药源植物,其根茎中薯蓣皂苷元含量居薯蓣属植物之冠,为我国的特有种。为了寻找高含量的资源、筛选新的生理活性成分,多年来我国学者做了大量的研究工作。主要概括了盾叶薯蓣的资源分布、薯蓣皂苷元的提取工艺、化学成分、药理、含量测定等方面的研究。     

The Rho kinase inhibitor Y-27632 facilitates the differentiation of bone marrow mesenchymal stem cells

The selective in vitro expansion and differentiation of multipotent stem cells are critical steps in cell-based regenerative therapies, but technical challenges have limited cell yield and thus the success of these potential treatments. The Rho GTPases and downstream Rho kinases (Rho coiled-coil kinases or ROCKs) are central regulators of cytoskeletal dynamics during the cell cycle and thus help determine the balance between stem cells self-renewal, lineage commitment, and apoptosis. Here, we examined if suppression of ROCK signaling enhances the efficacy of bone marrow-derived mesenchymal stem cells (BMSCs) differentiation into neurons and neuroglial cells. BMSCs were cultured in epidermal growth factor (EGF, 10 µg/l) and basic fibroblastic growth factor (bFGF, 10 µg/l) in the presence or absence of the Rho kinase inhibitor Y-27632 (10 µM). The expression levels of neuron-specific enolase (NSE) and glial fibrillary acidic protein (GFAP) were detected by immunofluorescence and Western blotting. The average number of NSE-positive cells increased from 83.20 ± 8.677 (positive ratio 0.2140 ± 0.0119) to 109.20 ± 8.430 (positive ratio 0.3193 ± 0.0161) per visual field in the presence of Y-27632, while GFAP-positive cell number increased from 96.30 ± 8.486 (positive ratio 0.18 ± 0.0152) to 107.50 ± 8.683 (positive ratio 0.27 ± 0.0115) (P < 0.05 for both). Both NSE and GFAP protein expression levels were enhanced significantly by Y-27632 treatment (NSE: 0.74 ± 0.05 vs. 1.03 ± 0.06; GFAP: 0.64 ± 0.08 vs. 0.97 ± 0.05, both P < 0.01) as indicated by Western blots. The Rho kinase inhibitor Y-27632 concomitant with EGF and bFGF stimulation promotes BMSC differentiation into neural cells. Control of Rho kinase activity may enhance the efficiency of stem cell-based treatments for neurodegenerative diseases.     

Insights into xanthine riboswitch structure and metal ion-mediated ligand recognition

Riboswitches are conserved functional domains in mRNA that mostly exist in bacteria. They regulate gene expression in response to varying concentrations of metabolites or metal ions. Recently, the NMT1 RNA motif has been identified to selectively bind xanthine and uric acid, respectively, both are involved in the metabolic pathway of purine degradation. Here, we report a crystal structure of this RNA bound to xanthine. Overall, the riboswitch exhibits a rod-like, continuously stacked fold composed of three stems and two internal junctions. The binding-pocket is determined by the highly conserved junctional sequence J1 between stem P1 and P2a, and engages a long-distance Watson–Crick base pair to junction J2. Xanthine inserts between a G–U pair from the major groove side and is sandwiched between base triples. Strikingly, a Mg²⁺ ion is inner-sphere coordinated to O6 of xanthine and a non-bridging oxygen of a backbone phosphate. Two further hydrated Mg²⁺ ions participate in extensive interactions between xanthine and the pocket. Our structure model is verified by ligand binding analysis to selected riboswitch mutants using isothermal titration calorimetry, and by fluorescence spectroscopic analysis of RNA folding using 2-aminopurine-modified variants. Together, our study highlights the principles of metal ion-mediated ligand recognition by the xanthine riboswitch.