丝状真菌代谢工程研究进展

丝状真菌(Filamentous fungi)作为重要的工业发酵微生物,在有机酸、蛋白质及次级代谢产物等关键生物基产品生产方面发挥着重要作用.自20世纪90年代代谢工程理念提出以来,尤其是代谢工程使能技术的创新及发展,极大地促进了丝状真菌细胞工厂的构建及其在工业发酵领域的应用.文中将系统介绍近年来丝状真菌代谢工程技术的...     

中国东北地区流动沙丘生态系统丘间低地地下芽库的时空变化

中国东北地区流动沙丘生态系统丘间低地地下芽库的时空变化 地下芽库在半干旱区沙丘生态系统植被恢复中起着重要作用。然而,目前针对流动沙丘丘间低地地下芽库时空变化的研究却很少。本研究通过调查一个生长季内5个不同面积流动沙丘丘间低地地下芽库的大小和组成,确定流动沙丘丘间低地地下芽库的时空变化。研究结果显示,中等面积丘间低地的总芽库密度与分蘖芽密度最高,茎基部芽密度呈相反趋势,而根茎芽密度不随丘间低地的面积变化而变化。地下芽库大小具有明显的季节变化特征。总芽密度在8月份达到高峰,10月份最低,根茎芽密度变化趋势与总芽密度相似,而茎基芽密度变化趋势与总芽密度相反,分蘖芽密度的变化不明显。以上结果表明,地下芽库密度随丘间低地的面积和季节变化而变化。这一结果有助于认识流动沙丘生态系统中植物生长的适应策略,并可为半干旱区沙丘植被恢复和保护提供理论指导。     

非磷酸化肌球蛋白在血小板衍生生长因子诱导的血管平滑肌细胞迁移中的作用

为了阐明非磷酸化肌球蛋白在平滑肌细胞迁移中的作用,研究探讨了非磷酸化肌球蛋白是否介导了血小板衍生生长因子(PDGF)诱导豚鼠脑基底动脉平滑肌细胞(GbaSM-4)的迁移。研究结果显示,20ng/ml以下剂量的PDGF可诱导GbaSM-4细胞发生迁移,此时肌球蛋白轻链(MLC20)磷酸化水平无变化。该迁移作用可被肌球蛋白特异性抑制剂blebbistatin所拮抗。应用RNA干扰技术抑制肌球蛋白轻链激酶表达,经免疫印迹检测经果显示,MLC20的磷酸化水平发生了显著下降;但对PDGF诱导的迁移作用无影响;在RNA干扰后blebbistatin也可抑制其迁移作用。体外ATP酶活性测定结果显示,blebbistatin对从平滑肌中提取的非磷酸化肌球蛋白的ATP酶活性有明显的抑制作用,其主要作用位点位于肌球蛋白头的头部S1。上述结果提示,非磷酸化的肌球蛋白参与了PDGF诱导的平滑肌细胞迁移。     

广西九里香根部的化学成分

从广西九里香根中分离到两个咔吧唑生物碱和一个甾醇,经波谱方法鉴定为九里香叶甲碱(1),九里香碱(2)和β-谷甾醇。     

The thymic stromal cell line MTSC4 induced thymocyte apoptosis in a non-MHC-restricted manner

Mouse thymic stromal cell line 4 (MTSC4) is one of the stromal cell lines established in our laboratory. While losing the characteristics of epithelial cells, they express some surface markers shared with thymic dendritic cells (TDCs). To further study the biological functions of these cells, we compared the capability of MTSC4 with TDCs in the induction of thymocyte apoptosis, using thymic reaggregation culture system. Apoptosis of thymocytes induced by MTSC4 and TDCs was measured by Annexin V and PI staining and analyzed by flow cytometry. We found that MTSC4 selectively augmented the apoptosis of CD4^ 8^ (DP) thymocytes. This effect was Fas/FasL independent and could not be blocked by antibodies to MHC class I and class II molecules. In addition, MTSC4 enhanced the apoptosis of DP thymocytes from different strains of mice, which implies that MTSC4-induced thymocyte apoptosis is not mediated by the TCR recognition of self peptide/MHC molecules. In contrast to MTSC4, thymocyte apoptosis induced by TDCs was MHC-restricted. Thus, MHC-independent fashion of stromal-DP thymocyte interaction may be one of the ways to induce thymocyte apoptosis in thymus. Our study has also shown that the interaction of MTSC4 stromal cells and thymocytes is required for the induction of thymocyte apoptosis.     

安徽农田表层土壤中有机氯农药的分布及其组成

以安徽省寿县等19个地区的农田表层土壤(0～20 cm)为对象,采用超声波提取,气相色谱法/电子捕获检测器(GC/ECD)检测,分析了α-六六六、β-六六六、γ-六六六、δ-六六六、op′-DDE、pp′-DDD、α-硫丹和百菌清等8种有机氯农药(OCPs)在农田表层土壤中的分布及组成特征.结果表明： 19个取样点中8种有机氯农药残留的总含量范围为ND～23.75 μg·kg^-1,其中PP′-DDD、γ-HCH为主要污染物,平均质量浓度分别为13.83和13.49 μg·kg^-1.与1990年的调查结果相比,六六六平均值含量呈明显下降趋势;与国内外土壤相比,安徽省农田表层土壤中的六六六(HCHs)处于中等偏高的水平.安徽省农田表层土壤中OCPs、HCHs和pp′-DDD平均浓度分别为48.58、28.64和13.83 μg·kg^-1,均未超过《土壤环境质量标准》(GB 15618-1995)的一级土壤质量标准(<50 μg·kg^-1),污染较轻.     

汉滩病毒M和S基因不同拼接方式嵌合基因免疫效果的研究

本文在前期工作的基础上,构建了汉滩病毒76－118株M基因G2片段与S基因5'端0.7Kb片段的嵌合基因真核表达载体pcDNA3.1-G2S0.7及pcDNA3.1-S0．7G2;用该质粒免疫BALB/c小鼠,结果表明两种质粒免疫小鼠可同时诱导产生抗滩滩病核蛋白（NP)及糖蛋白（GP）特异性的抗体,且前者刺激产生的抗体效价明显高于后者。淋巴细胞增殖实验表明,pcDNA3.1-G2S0.7组免疫小鼠脾细胞时NP及GP的增殖指数均明显高于空载体对照组,而pcDNA3.1-S0．7G2组未检测到其淋巴细胞有明显的增殖。这说明汉滩病毒M基因G2片段及S基因0.7Kb片段的嵌合基因既可刺激机体产生特异的抗汉滩病毒体液免疫应答,也可刺激机体产生特异的细胞免疫应答。不同拼接方式对嵌合基因免疫效果有很大影响,嵌合基因G2S0．7这种拼接方式明显优于S0．7G2。     

Proteomics analysis of serum from mutant mice reveals lysosomal proteins selectively transported by each of the two mannose 6-phosphate receptors

Most mammalian cells contain two types of mannose 6-phosphate (Man-6-P) receptors (MPRs): the 300 kDa cation-independent (CI) MPR and 46 kDa cation-dependent (CD) MPR. The two MPRs have overlapping function in intracellular targeting of newly synthesized lysosomal proteins, but both are required for efficient targeting. Despite extensive investigation, the relative roles and specialized functions of each MPR in targeting of specific proteins remain questions of fundamental interest. One possibility is that most Man-6-P glycoproteins are transported by both MPRs, but there may be subsets that are preferentially transported by each. To investigate this, we have conducted a proteomics analysis of serum from mice lacking either MPR with the reasoning that lysosomal proteins that are selectively transported by a given MPR should be preferentially secreted into the bloodstream in its absence. We purified and identified Man-6-P glycoproteins and glycopeptides from wild-type, CDMPR-deficient, and CIMPR-deficient mouse serum and found both lysosomal proteins and proteins not currently thought to have lysosomal function. Different mass spectrometric approaches (spectral count analysis of nanospray LC-MS/MS experiments on unlabeled samples and LC-MALDI/TOF/TOF experiments on iTRAQ-labeled samples) revealed a number of proteins that appear specifically elevated in serum from each MPR-deficient mouse. Man-6-P glycoforms of cellular repressor of E1A-stimulated genes 1, tripeptidyl peptidase I, and heparanase were elevated in absence of the CDMPR and Man-6-P glycoforms of alpha-mannosidase B1, cathepsin D, and prosaposin were elevated in the absence of the CIMPR. Results were confirmed by Western blot analyses for select proteins. This study provides a comparison of different quantitative mass spectrometric approaches and provides the first report of proteins whose cellular targeting appears to be MPR-selective under physiological conditions.     

硫酸盐还原菌固态发酵产品酶活测定方法的建立

为建立生物质固载硫酸盐还原菌（SRB）产品酶活的检测方法,首先确定亚硫酸盐还原酶（SiR）作为SRB胞外酶表征其生物活性的可行性;其次对固态发酵产品制备粗酶液的各种条件进行探讨,研究浸泡介质、浸泡时间、超声功率、超声时间、预处理方式等单因素对酶活的影响;由正交实验确定粗酶液制备的最佳工况。实验结果表明,先浸提后超声处理的酶活值显著高于仅用超声、浸提的结果。为确保酶活值测定不受干扰,排除超声温度对结果的影响,确定实验最佳工况：2g固态发酵产品经磷酸盐缓冲液25mL、浸提60min,酶活值为1．0799U／g。生物质固载SRB酶活检测方法的建立为固态发酵产品活性评价、固态发酵条件优化、可渗透反应墙生化性能评估提供一定的理论依据。     

miR-22 regulates C2C12 myoblast proliferation and differentiation by targeting TGFBR1

Recently, miR-22 was found to be differentially expressed in different skeletal muscle growth period, indicated that it might have function in skeletal muscle myogenesis. In this study, we found that the expression of miR-22 was the most in skeletal muscle and was gradually up-regulated during mouse myoblast cell (C2C12 myoblast cell line) differentiation. Overexpression of miR-22 repressed C2C12 myoblast proliferation and promoted myoblast differentiation into myotubes, whereas inhibition of miR-22 showed the opposite results. During myogenesis, we predicted and verified transforming growth factor beta receptor 1 (TGFBR1), a key receptor of the TGF-β/Smad signaling pathway, was a target gene of miR-22. Then, we found miR-22 could regulate the expression of TGFBR1 and down-regulate the Smad3 signaling pathway. Knockdown of TGFBR1 by siRNA suppressed the proliferation of C2C12 cells but induced its differentiation. Conversely, overexpression of TGFBR1 significantly promoted proliferation but inhibited differentiation of the myoblast. Additionally, when C2C12 cells were treated with different concentrations of transforming growth factor beta 1 (TGF-β1), the level of miR-22 in C2C12 cells was reduced. The TGFBR1 protein level was significantly elevated in C2C12 cells treated with TGF-β1. Moreover, miR-22 was able to inhibit TGF-β1-induced TGFBR1 expression in C2C12 cells. Altogether, we demonstrated that TGF-β1 inhibited miR-22 expression in C2C12 cells and miR-22 regulated C2C12 cell myogenesis by targeting TGFBR1.