不同水分胁迫对刺槐生理的影响

本文以二年生刺槐为供试材料,探讨水分胁迫对其生理的影响,为园林绿地中刺槐的灌溉提供理论指导.结果表明,水淹和干旱胁迫明显影响刺槐的生长.水淹11 d后,qP值、ETR值、Fv'/Fm'值和净光合速率的明显下降,qN值上升,19 d后植株死亡.中度干旱胁迫(土壤含水量在8%～15%之间)对植株生长有一定影响,表现为净光合速率的下降和qN值上升等;干旱胁迫(土壤含水量低于10%)严重抑制植株的生长,表现为qN值上升,而qP、ETR、Fv'/Fm'和净光合速率下降,处理19 d后干旱组植株死亡.轻度干旱(土壤含水量在15%～25%之间)适宜植物生长,表现为3次测定的qP、ETR、Fv'/Fm'、Fv/Fm和净光合速率都较高且稳定.土壤含水量日变化在15%～25%以内受轻度干旱胁迫是园林中刺槐的最佳灌溉方式,既不影响景观效果,同时也能节约灌溉用水.     

Mite allergen Der-p2 triggers human B lymphocyte activation and Toll-like receptor-4 induction

Background

Allergic disease can be characterized as manifestations of an exaggerated inflammatory response to environmental allergens triggers. Mite allergen Der-p2 is one of the major allergens of the house dust mite, which contributes to TLR4 expression and function in B cells in allergic patients. However, the precise mechanisms of Der-p2 on B cells remain obscure.

Methodology/Principal Findings

We investigated the effects of Der-p2 on proinflammatory cytokines responses and Toll-like receptor-4 (TLR4)-related signaling in human B cells activation. We demonstrated that Der-p2 activates pro-inflammatory cytokines, TLR4 and its co-receptor MD2. ERK inhibitor PD98059 significantly enhanced TLR4/MD2 expression in Der-p2-treated B cells. Der-p2 markedly activated mitogen-activated protein kinase (MAPK) phosphatase-1 (MKP-1) and decreased p38 phosphorylation in B cells. MKP-1-siRNA downregulated TLR4/MD2 expression in Der-p2-treated B cells. In addition, Der-p2 significantly up-regulated expression of co-stimulatory molecules and increased B cell proliferation. Neutralizing Der-p2 antibody could effectively abrogate the Der-p2-induced B cell proliferation. Der-p2 could also markedly induce NF-κB activation in B cells, which could be counteracted by dexamethasone.

Conclusions/Significance

These results strongly suggest that Der-p2 is capable of triggering B cell activation and MKP-1-activated p38/MAPK dephosphorylation-regulated TLR4 induction, which subsequently enhances host immune, defense responses and development of effective allergic disease therapeutics in B cells.     

Polyclonal Antibodies in Microplates to Predict the Maximum Adsorption Activities of Enzyme/Mutants from Cell Lysates

With microplate-immobilized polyclonal antibodies against a starting enzyme or its active mutant bearing consistent accessible epitopes, the maximum activity of an adsorbed enzyme/mutant (Vs) was predicted for comparison to recognize weakly-positive mutants. Rabbit antisera against Escherichia coli alkaline phosphatase (ECAP) were fractionated with 33% ammonium sulfate to yield crude polyclonal antibodies for conventional immobilization in 96-well microplates. The response curve of the activities of ECAP/mutant adsorbed by the immobilized polyclonal antibodies to protein quantities from a cell lysate was fit to an approximation model to predict Vs. With 0.4 μg crude polyclonal antibody for immobilization, Vs was consistent for ECAP in cell lysates bearing fourfold differences in its apparent specific activities when its abundance was greater than 0.9%. The ratio of Vs of the mutant R168K to that of ECAP was 1.5?±?0.1 (n?=?2), consistent with that of their specific activities after affinity purification. Unfortunately, the prediction of Vs with polyclonal antibodies that saturated microplate wells was ineffective to Pseudomonas aeruginosa arylsulfatase bearing less than 2% specific activity of ECAP. Therefore, with microplate-immobilized polyclonal antibodies to adsorb enzyme/mutants from cell lysates, high-throughput prediction of Vs was practical to recognize weakly-positive mutants of starting enzymes bearing fairly-high activities.     

Ganocapenoids A–D: Four new aromatic meroterpenoids from Ganoderma capense

Four new aromatic meroterpenoids, ganocapenoids A–D (1–4), together with twelve known analogues (5–16) were isolated from the fruiting bodies of Ganoderma capense. The structures of new compounds were determined through spectroscopic methods including 1D and 2D NMR and MS analyses. Their absolute configurations were assigned by ECD calculations and specific rotation comparison. The biological activities of these substances toward regulation of lipid metabolism, neurite outgrowth-promoting activity, and AchE inhibition were assessed. Compound 15 was found to be able to block lipid accumulation at a concentration of 20?μM, and compounds 4a, 4b, and 11 show moderate neurite outgrowth-promoting activity at 10?μM, while compounds 3, 6, 11, and 13 exhibit potent AchE inhibition with the IC₅₀ values of 28.6?±?1.9, 18.7?±?1.6, 8.2?±?0.2, 26.0?±?2.9?μM, respectively.     

A new geranylgeranyl diphosphate synthase gene from Ginkgo biloba, which intermediates the biosynthesis of the key precursor for ginkgolides.

Geranylgeranyl diphosphate synthase (GGPPS, EC: 2.5.1.29) catalyzes the biosynthesis of geranylgeranyl diphosphate (GGPP), which is a key precursor for ginkgolide biosynthesis. Here we reported for the first time the cloning of a new full-length cDNA encoding GGPPS from the living fossil plant Ginkgo biloba. The full-length cDNA encoding G. biloba GGPPS (designated as GbGGPPS) was 1657bp long and contained a 1176bp open reading frame encoding a 391 amino acid protein. Comparative analysis showed that GbGGPPS possessed a 79 amino acid transit peptide at its N-terminal, which directed GbGGPPS to target to the plastids. Bioinformatic analysis revealed that GbGGPPS was a member of polyprenyltransferases with two highly conserved aspartate-rich motifs like other plant GGPPSs. Phylogenetic tree analysis indicated that plant GGPPSs could be classified into two groups, angiosperm and gymnosperm GGPPSs, while GbGGPPS had closer relationship with gymnosperm plant GGPPSs.     

A smooth muscle Cav1.2 calcium channel splice variant underlies hyperpolarized window current and enhanced state-dependent inhibition by nifedipine

Native smooth muscle L-type Ca(v)1.2 calcium channels have been shown to support a fraction of Ca(2+) currents with a window current that is close to resting potential. The smooth muscle L-type Ca(2+) channels are also more susceptible to inhibition by dihydropyridines (DHPs) than the cardiac channels. It was hypothesized that smooth muscle Ca(v)1.2 channels exhibiting hyperpolarized shift in steady-state inactivation would contribute to larger inhibition by DHP, in addition to structural differences of the channels generated by alternative splicing that modulate DHP sensitivities. In addition, it has also been shown that alternative splicing modulates DHP sensitivities by generating structural differences in the Ca(v)1.2 channels. Here, we report a smooth muscle L-type Ca(v)1.2 calcium channel splice variant, Ca(v)1.2SM (1/8/9(*)/32/Delta33), that when expressed in HEK 293 cells display hyperpolarized shifts for steady-state inactivation and activation potentials when compared with the established Ca(v)1.2b clone (1/8/9(*)/32/33). This variant activates from more negative potentials and generates a window current closer to resting membrane potential. We also identified the predominant cardiac isoform Ca(v)1.2CM clone (1a/8a/Delta9(*)/32/33) that is different from the established Ca(v)1.2a (1a/8a/Delta9(*)/31/33). Importantly, Ca(v)1.2SM channels were shown to be more sensitive to nifedipine blockade than Ca(v)1.2b and cardiac Ca(v)1.2CM channels when currents were recorded in either 5 mM Ba(2+) or 1.8 mM Ca(2+) external solutions. This is the first time that a smooth muscle Ca(v)1.2 splice variant has been identified functionally to possess biophysical property that can be linked to enhanced state-dependent block by DHP.     

Nobiletin, a citrus flavonoid, suppresses invasion and migration involving FAK/PI3K/Akt and small GTPase signals in human gastric adenocarcinoma AGS cells

Nobiletin, a compound isolated from citrus fruits, is a polymethoxylated flavone derivative shown to have anti-inflammatory, antitumor, and neuroprotective properties. This study has investigated that nobiletin exerted inhibitory effects on the cell adhesion, invasion, and migration abilities of a highly metastatic AGS cells under non-cytotoxic concentrations. Data also showed nobiletin could inhibit the activation of focal adhesion kinase (FAK) and phosphoinositide-3-kinase/Akt (PI3K/Akt) involved in the downregulation of the enzyme activities, protein expressions, messenger RNA levels of matrix metalloproteinase-2 (MMP-2), and matrix metalloproteinase-2 (MMP-9). Also, our data revealed that nobiletin inhibited FAK/PI3K/Akt with concurrent reduction in the protein expressions of Ras, c-Raf, Rac-1, Cdc42, and RhoA by western blotting, whereas the protein level of RhoB increased progressively. Otherwise, nobiletin-treated AGS cells showed tremendously decreased in the phosphorylation and degradation of inhibitor of kappaBα (IκBα), the nuclear level of NF-κB, and the binding ability of NF-κB to NF-κB response element. Furthermore, nobiletin significantly decreased the levels of phospho-Akt and MMP-2/9 in Akt1-cDNA-transfected cells concomitantly with a marked reduction in cell invasion and migration. These results suggest that nobiletin can reduce invasion and migration of AGS cells, and such a characteristic may be of great value in the development of a potential cancer therapy.     

双向BN/SDS-PAGE分离鉴定痢疾杆菌福氏5型M90T株毒力相关膜蛋白复合物

通过蓝色非变性凝胶电泳(BN-PAGE)比较痢疾杆菌福氏5型野生株M90T和大质粒缺失株M90T△T的膜蛋白复合物,发现一个野生株特有的复合物,其分子量的为290 kD,命名为M90T-290.通过第二向SDS-PAGE分离M90T-290得到6个蛋白亚基,质谱鉴定为:一个由大质粒编码的毒力蛋白Apyrase(ATP-二磷酸水解酶)和5个染色体编码的蛋白,这些蛋白可能以膜复合物的形式影响毒力蛋白IcsA的单极性分布和痢疾杆菌在细胞间的扩散.这个新发现的毒力相关膜复合物在痢疾杆菌的致病过程中可能发挥重要作用.     

Phylogeographic analysis reveals two cryptic species of the endangered fern <Emphasis Type="Italic">Ceratopteris</Emphasis><Emphasis Type="Italic">thalictroides</Emphasis> (L.) Brongn. (Parkeriaceae) in China

Ceratopteris thalictroides (L.) Brongn. (Parkeriaceae) is a difficult fern species to taxonomically classify. Three cryptic species were revealed in the previous studies, referred to as the north type, the south type, and the third type. Because much of the distribution range of C. thalictroides in China was not included in the sampling of the previous studies, the taxonomic complexity of C. thalictroides in China remained uncertain. In order to identify the uncharacterized cryptic species, we examined four chloroplast DNA (cpDNA) non-coding regions and compared sequence variation within this species complex. Sequence data were obtained from 143 individuals in 24 populations throughout the natural distribution of the species in China. Nineteen haplotypes were identified. Molecular systematic and phylogeographical analyses revealed two genetically distinct clusters of cpDNA haplotypes in China. One cluster included haplotypes associated with the north type, and another with the south type cryptic species. The N _ST value was significantly higher than the G _ST value (N _ST = 0.768 > G _ST = 0.434, P < 0.05), indicating the presence of a significant phylogeographical structure of C. thalictroides in China. The results of AMOVA analysis showed a significant inter-group differentiation (F _ST = 0.918; P < 0.001). Analyses based on different, but complementary methods suggest that in China, C. thalictroides contains only two of the cryptic species (the north and south types). Two haplotypes, H8 and H17, of the interior node in the minimum-spanning network (MSN) of cpDNA haplotypes are widespread. The origin of the widespread haplotypes in China may have resulted from long-distance dispersal to China.     

Identification of histone demethylases in Saccharomyces cerevisiae

Based on the prediction that histone lysine demethylases may contain the JmjC domain, we examined the methylation patterns of five knock-out strains (ecm5Delta, gis1Delta, rph1Delta, jhd1Delta, and jhd2Delta (yjr119cDelta)) of Saccharomyces cerevisiae. Mass spectrometry (MS) analyses of histone H3 showed increased modifications in all mutants except ecm5Delta. High-resolution MS was used to unequivocally differentiate trimethylation from acetylation in various tryptic fragments. The relative abundance of specific fragments indicated that histones K36me3 and K4me3 accumulate in rph1Delta and jhd2Delta strains, respectively, whereas both histone K36me2 and K36me accumulate in gis1Delta and jhd1Delta strains. Analyses performed with strains overexpressing the JmjC proteins yielded changes in methylation patterns that were the reverse of those obtained in the complementary knock-out strains. In vitro enzymatic assays confirmed that the JmjC domain of Rph1 specifically demethylates K36me3 primarily and K36me2 secondarily. Overexpression of RPH1 generated a growth defect in response to UV irradiation. The demethylase activity of Rph1 is responsible for the phenotype. Collectively, in addition to Jhd1, our results identified three novel JmjC domain-containing histone demethylases and their sites of action in budding yeast S. cerevisiae. Furthermore, the methodology described here will be useful for identifying histone demethylases and their target sites in other organisms.