大豆过氧化物酶在毕赤酵母中功能表达

【目的】大豆过氧化物酶(SBP)作用底物广泛、比活高、热稳定性好,使其在免疫检测、工业污染废水处理领域有着广泛的应用潜力。现有的生产方法主要是从大豆壳中提取,这种方法产量低,成本高,远不能满足于工业应用要求,本研究希望实现在毕赤酵母中高效表达有功能活性的大豆过氧化物酶。【方法】将大豆过氧化物酶基因以及C末端截短20个氨基酸的基因克隆pPIC-9K载体中,并在毕赤酵母X-33中诱导表达。同时还将糖基化位点的天冬酰胺突变成为谷氨酰胺,研究糖基化位点对表达的影响。【结果】全长SBP在毕赤酵母中表达是无活性的,只有截短的SBP△20在试管发酵的表达活力达23.5 U/mL,经过糖基化位点的突变表明130、144、185、197对酶活非常重要,不能突变;211和216位点去糖基化突变对酶活有所提高。【结论】经过发酵条件的优化,在5 L的发酵罐中发酵液上清最高酶活力达510 U/mL,是目前报道的最高水平。     

中国虫草一新记录种

【目的】对一个寄生鳞翅目幼虫的虫草标本Dxhir140901进行分类鉴定。【方法】采用形态学比较和基于ITS1-5.8S-ITS2rDNA的系统发育与进化网络分析进行鉴定。【结果】形态学观察：标本的分离菌株形态显示其为典型的被毛孢属真菌,具有两型产孢结构：A型产孢细胞柱状,(1.8?6.3) μm×1.8 μm;B型产孢细胞锥形,基部柱状,向上逐渐变细无明显颈部,基部宽3?3.8 μm,长21?63 μm,颈部宽1.8?2.0 μm,菌丝末端可直接形成产孢细胞;孢子橘瓣形或卵形,(8.1?10.8) μm×(2.7?5.4) μm,具粘液,黏液层厚1.8?2.7 μm。系统发育分析结果显示该菌株与巨针线形虫草Ophiocordyceps macroacicularis聚为一支,支持率为98%,进化网络分析也支持上述结果。【结论】通过与O. macroacicularis的形态比较和分子系统学分析结果,Dxhir140901及其分离株Gzuifr-hir140901为巨针线形虫草Ophiocordyceps macroacicularis S. Ban, T. Sakane & Nakagiri的无性阶段,该种为中国新记录种。     

城乡建设用地关联对生态系统服务的影响——以洞庭湖地区为例

城乡建设用地数量及相互联系的变化是空间要素及结构演化的重要方面,深刻影响着生态环境质量和生态系统服务水平,研究城乡建设用地关联对生态系统服务的影响对国土空间高质量发展具有重要意义。以洞庭湖地区为研究对象,从土地利用关联视角出发,运用引力模型、InVEST模型、面板数据模型等方法,定量研究了城乡建设用地关联对生态系统服务的影响。结果表明：（1）洞庭湖地区城镇建设用地规模呈现上升趋势,农村居民点用地规模呈现下降趋势;（2）城乡建设用地关联强度呈不断增强趋势,且强关联网格以建成区为核心向周边扩展,低关联网格分布于远离建成区的地方;（3）生境质量和碳储量呈现下降趋势,食物供给则呈现上升趋势,各项生态系统服务价值具有显著的空间异质性;（4）城乡建设用地关联强度对生态系统服务具有负向影响,人口密度、GDP密度、夜间灯光、道路网络对生态系统服务也呈现负向影响,归一化差异植被指数与生态系统服务之间则存在显著的正相关。研究结果有助于了解关联视角下城乡建设用地对生态系统服务的影响机制,可以为制定合理的城乡空间发展战略提供科学依据。     

3,3′,4,5′-Tetramethoxy-trans-stilbene Improves Insulin Resistance by Activating the IRS/PI3K/Akt Pathway and Inhibiting Oxidative Stress

The potential anti-diabetic effect of resveratrol derivative, 3,3′,4,5′-tetramethoxy-trans-stilbene (3,3′,4,5′-TMS) and its underlying mechanism in high glucose (HG) and dexamethasone (DXMS)-stimulated insulin-resistant HepG2 cells (IR-HepG2) were investigated. 3,3′,4,5′-TMS did not reduce the cell viability of IR-HepG2 cells at the concentrations of 0.5–10 µM. 3,3′,4,5′-TMS increased the potential of glucose consumption and glycogen synthesis in a concentration-dependent manner in IR-HepG2 cells. 3,3′,4,5′-TMS ameliorated insulin resistance by enhancing the phosphorylation of glycogen synthase kinase 3 beta (GSK3β), inhibiting phosphorylation of insulin receptor substrate-1 (IRS-1), and activating phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) pathway in IR-HepG2 cells. Furthermore, 3,3′,4,5′-TMS significantly suppressed levels of reactive oxygen species (ROS) with up-regulation of nuclear factor erythroid 2-related factor 2 (Nrf2) expression. To conclude, the beneficial effect of 3,3′,4,5′-TMS against insulin resistance to increase glucose consumption and glycogen synthesis was mediated through activation of IRS/PI3K/Akt signaling pathways in the IR-HepG2 cells, accomplished with anti-oxidative activity through up-regulation of Nrf2.     

SARS-CoV-2 envelope protein causes acute respiratory distress syndrome (ARDS)-like pathological damages and constitutes an antiviral target

Cytokine storm and multi-organ failure are the main causes of SARS-CoV-2-related death. However, the origin of excessive damages caused by SARS-CoV-2 remains largely unknown. Here we show that the SARS-CoV-2 envelope (2-E) protein alone is able to cause acute respiratory distress syndrome (ARDS)-like damages in vitro and in vivo. 2-E proteins were found to form a type of pH-sensitive cation channels in bilayer lipid membranes. As observed in SARS-CoV-2-infected cells, heterologous expression of 2-E channels induced rapid cell death in various susceptible cell types and robust secretion of cytokines and chemokines in macrophages. Intravenous administration of purified 2-E protein into mice caused ARDS-like pathological damages in lung and spleen. A dominant negative mutation lowering 2-E channel activity attenuated cell death and SARS-CoV-2 production. Newly identified channel inhibitors exhibited potent anti-SARS-CoV-2 activity and excellent cell protective activity in vitro and these activities were positively correlated with inhibition of 2-E channel. Importantly, prophylactic and therapeutic administration of the channel inhibitor effectively reduced both the viral load and secretion of inflammation cytokines in lungs of SARS-CoV-2-infected transgenic mice expressing human angiotensin-converting enzyme 2 (hACE-2). Our study supports that 2-E is a promising drug target against SARS-CoV-2.Subject terms: Cell death, Molecular biology     

TEB/POLQ plays dual roles in protecting Arabidopsis from NO-induced DNA damage

Nitric oxide (NO) is a key player in numerous physiological processes. Excessive NO induces DNA damage, but how plants respond to this damage remains unclear. We screened and identified an Arabidopsis NO hypersensitive mutant and found it to be allelic to TEBICHI/POLQ, encoding DNA polymerase θ. The teb mutant plants were preferentially sensitive to NO- and its derivative peroxynitrite-induced DNA damage and subsequent double-strand breaks (DSBs). Inactivation of TEB caused the accumulation of spontaneous DSBs largely attributed to endogenous NO and was synergistic to DSB repair pathway mutations with respect to growth. These effects were manifested in the presence of NO-inducing agents and relieved by NO scavengers. NO induced G2/M cell cycle arrest in the teb mutant, indicative of stalled replication forks. Genetic analyses indicate that Polθ is required for translesion DNA synthesis across NO-induced lesions, but not oxidation-induced lesions. Whole-genome sequencing revealed that Polθ bypasses NO-induced base adducts in an error-free manner and generates mutations characteristic of Polθ-mediated end joining. Our experimental data collectively suggests that Polθ plays dual roles in protecting plants from NO-induced DNA damage. Since Polθ is conserved in higher eukaryotes, mammalian Polθ may also be required for balancing NO physiological signaling and genotoxicity.     

半干旱区湿地-干草原交错带边界判定及其变化

鉴于目前国内对于半干旱区及小尺度上湿地-干草原交错带边界界定的研究较少,利用交错带对其周围环境变化具有提前指示作用的原理,以宁夏盐池四儿滩湿地为例,首先探讨游动分割窗技术在半干旱区湿地-干草原交错带边界判定中的可行性,其次再对2006年—2010年湿地-干草原交错带的宽度和边界进行判定,通过交错带的宽度和边界变化并结合交错带内植被的变化情况,综合对整个湿地生态系统的健康状况和湿地退化进行研究。结果表明:研究的尺度大小以及交错带过渡性是否明显将决定适宜窗口宽度的大小,这点在选择最小适宜窗口宽度时体现更为显著;降雨量是决定该区交错带的位置与宽度的最主要因素,但降雨量并不是决定该区交错带内植被变化状况的最主要因素,交错带距湿地核心区的距离是影响交错带内物种丰富度指数变化的最主要因素;由于保护区的成立,四儿滩湿地在2006年—2007年间,交错带宽度变大,湿地状况转好,2008年—2010年,由于铁路建设,湿地遭到破坏,交错带萎缩,湿地出现退化现象。湿地-干草原交错带作为一个敏感地带,它的变化情况可以对整个湿地生态系统的变化情况作出一定的提前预示;在半干旱区湿地的健康评价和退化研究中,通过交错带的变化来反映整个湿地生态系统的健康情况是一种可行的思路和方法。