MicroRNAs and their role in viral infection

Recently, a class of about 22 nucleotides (nt) small RNA has been discovered in many eukaryotes, termed microRNAs (miRNAs), which have a variety of functions. Many recent findings have demonstrated that viruses can also encode their own miRNAs. Meanwhile, other findings reveal a relationship between host miRNA and viral infection. These findings suggest a tight relationship between host and viral infection via miRNA pathway. This article introduces the miRNAs encoded by viruses and reviews the advances of the interaction of the mammalian host miRNAs and viral infection.     

Gene conjugation of molecular adjuvant C3d3 to hCGbeta increased the anti-hCGbeta Th2 and humoral immune response in DNA immunization

Human chorionic gonadotropin (hCG) has been used as an anti-fertility vaccine and as a target for cancer immunotherapy. We have explored the use of three copies of C3d in DNA vaccine as molecular adjuvant to improve the immunogenicity of this hormone in previous work and found that the immune response induced by pcDNA3-hCGbeta-C3d3 has been enhanced 243-fold compared with pcDNA3-hCGbeta following DNA immunization in BALB/c mice. In the present study, a new functionally active DNA vaccine of hCGbeta-C3d3 chimera based on pCMV4 vector has been described. We compared the expression efficiency of pCMV4 and pcDNA3 eukaryotic vectors for hCGbeta and hCGbeta-C3d3 fusion protein and the immune response of mice immunized with pcDNA3-hCGbeta, pCMV4-hCGbeta, pcDNA3-hCGbeta-C3d3 and pCMV4-hCGbeta-C3d3, respectively, at 25, 50 and 100 pmol dose, and further analyzed the levels of Th1 and Th2 cytokines produced by spleen lymphocytes of the immunized mice upon hCG restimulation in vitro. It was found that pCMV4 vector achieved 1.3-1.5-fold higher protein expression and raised 1.1-1.2 (primary) and 1.2-1.3 (booster) logs higher titer of anti-hCGbeta IgG than pcDNA3. Mice vaccinated with 50 pmol of hCGbeta-C3d3-DNAs elicited the highest titer of hCGbeta-specific antibody among the serial doses and the immune response induced by pCMV4-hCGbeta-C3d3 were, respectively, 1.3, 1.3 and 1.2 logs higher than that of pcDNA3-hCGbeta-C3d3 and 2.2, 2.9 and 2.4 logs higher than that of pCMV4-hCGbeta at week 2 following the booster immunization. Moreover, we observed that the production of IL-4 and IL-10 increased in mice vaccinated with hCGbeta-C3d3-DNAs and the ratio of IL-4/IFN-(gamma) showed a Th2 bias of immune response in the mice immunized with hCGbeta-C3d3-DNAs. These findings indicated that gene fusion of C3d3 to hCGbeta, as a means of harnessing the adjuvant potential of the innate immune system, may improve the antigen-specific Th2 humoral immune response of the hCGbeta DNA vaccine and the pCMV4 vector is a more ideal eukaryotic vector for DNA vaccine than pcDNA3.     

MST1 promotes apoptosis through phosphorylation of histone H2AX

MST1 (mammalian STE20-like kinase 1) is a serine/threonine kinase that is cleaved and activated by caspases during apoptosis. Overexpression of MST1 induces apoptotic morphological changes such as chromatin condensation, but the mechanism is not clear. Here we show that MST1 induces apoptotic chromatin condensation through its phosphorylation of histone H2AX at Ser-139. During etoposide-induced apoptosis in Jurkat cells, the cleavage of MST1 directly corresponded with strong H2AX phosphorylation. In vitro kinase assay results showed that MST1 strongly phosphorylates histone H2AX. Western blot and kinase assay results with a mutant S139A H2AX confirmed that MST1 phosphorylates H2AX at Ser-139. Direct binding of MST1 and H2AX can be detected when co-expressed in HEK293 cells and was also confirmed by an endogenous immunoprecipitation study. When overexpressed in HeLa cells, both the MST1 full-length protein and the MST1 kinase domain (MST1-NT), but not the kinase-negative mutant (MST1-NT-KN), could induce obvious endogenous histone H2AX phosphorylation. The caspase-3 inhibitor benzyloxycarbonyl-DEVD-fluoromethyl ketone (Z-DEVD-fmk) attenuates phosphorylation of H2AX by MST1 but cannot inhibit MST1-NT-induced histone H2AX phosphorylation, indicating that cleaved MST1 is responsible for H2AX phosphorylation during apoptosis. Histone H2AX phosphorylation and DNA fragmentation were suppressed in MST1 knockdown Jurkat cells after etoposide treatment. Taken together, our data indicated that H2AX is a substrate of MST1, which functions to induce apoptotic chromatin condensation and DNA fragmentation.     

家蚕二分浓核病毒VD1-ORF4基因截短多肽的表达和抗体制备

通过PCR扩增家蚕二分浓核病毒(Bombyx mori bidensovirus,BmBDV)VD1-ORF4基因序列中的两个DNA片段,将测序正确的两个目的片段分别亚克隆到原核表达载体pET-30a上,通过不同浓度的IPTG对含有重组质粒的大肠杆菌BL21(DE3)进行诱导,对诱导产物进行SDS-PAGE和Westen blot分析.结果表明,这两个截短多肽都获得了表达,其N-端融合有6个组氨酸.将割胶纯化的蛋白多肽与佐剂充分研磨,以研磨后的匀浆液对昆明小鼠进行皮下多点注射.获得的抗血清分别对原核诱导表达产物进行Western blot分析,结果表明,在特定的位置都能杂交到一条特异的蛋白带,表明制备的两个多抗能为深入研究VD1-ORF4基因的功能提供基础.     

不同盐碱类型胁迫对红地球/贝达葡萄植株离子分布的影响

以盆栽红地球/贝达葡萄为试材,定量浇灌NaCl、Na₂SO₄、NaHCO₃、NH₄Cl和(NH₄)₂SO₄,筛选导致葡萄叶片黄化的盐、碱离子,研究不同盐碱类型胁迫对葡萄植株离子分布的影响.结果表明： NaHCO₃对植株影响最大,叶片在处理14 d时出现黄化症状,而NaCl和NH₄Cl处理28 d时出现黄化症状.NaHCO₃和NaCl处理均显著增加了植株各器官中Na^+含量,NaHCO₃处理根中Na⁺含量是对照的 6.4倍;这两种盐处理均降低了除叶片外其他器官中的K⁺含量,NaHCO₃处理显著降低了各器官中K/Na,根中K/Na仅为0.1,NaCl处理降低了除茎外其他器官中K/Na;这两种盐处理还降低了Ca²⁺、Mg²⁺、Fe²⁺向地上部的运输.NH₄Cl、(NH₄)₂SO₄和Na₂SO₄处理降低了植株各器官中K/Na,以NH₄Cl处理显著.碱性盐NaHCO₃对葡萄叶片黄化影响最大,其次是中性盐NaCl,再次是NH₄Cl,而(NH₄)₂SO₄和Na₂SO₄影响较小.     

广西眼镜蛇蛇毒磷脂酶A_2的分离纯化及其对HSC-T6细胞的作用

目的对广西眼镜蛇毒中磷脂酶A2(PLA2)进行分离纯化,测定其对肝星状细胞HSC-T6的增殖抑制作用。方法采用Sephadex G-50凝胶层析柱、CM-Sepharose CL-6B离子交换柱、Macro-prep High S预装柱结合的方法分离广西眼镜蛇粗毒,经平板法测定各峰的PLA2活性;经SDS-PAGE电泳鉴定终产物纯度并测定分子量,NanoLC-ESI-MS/MS鉴定其组分;CCK-8法测定PLA2对肝星状细胞(HSC-T6)的增殖抑制作用,确定其凋亡的最小毒性浓度。结果 Sephadex G-50凝胶层析柱、CM-Sepharose CL-6B离子交换柱、Macro-prep High S预装柱层析法,得到第Ⅲ峰具PLA2活性,且达到电泳纯,经NanoLCESI-MS/MS鉴定其为PLA2,分子量约为14.06kD;PLA2在0~1μg/ml的浓度下对HSC-T6细胞具有一定的促增殖作用,2μg/ml时细胞数达到最大值,4~16μg/ml时对细胞生长有抑制作用,且随浓度增大细胞数降低。结论采用Sephadex G-50、CM-Sepharose CL-6B、Macro-prep High S预装柱结合的方法对广西眼镜蛇毒进行分离纯化,得到电泳纯且具PLA2活性的磷脂酶A2;广西眼镜蛇毒PLA2对肝星状细胞HSC-T6增殖有抑制作用,PLA2对HSC-T6细胞的最小毒性浓度为2μg/ml。     

湖南省双季稻生产系统碳效率

提高农作物生产系统的碳效率是实现低碳农业的重要途径之一.本文采用2004—2012年农作物产量、农田作物生产系统农资投入等统计数据,利用生命周期法和投入产出法,对湖南省双季稻生产系统碳排放、碳吸收和碳效率特征及其动态进行估算.结果表明:湖南省2004—2012年双季稻生产系统年均碳排放总量为656.4×107kg CE,其中化肥和农药生产运输碳排放占农资投入碳排放总量的大部分,分别约占70.0%和15.9%,碳排放总量年际间持续降低,年均降低率为2.4%,碳排放强度则表现为增长趋势;湖南双季稻生产系统2004—2012年年均碳吸收总量为1547.0×107kg C,也呈现逐年降低的趋势,年降低率为1.2%,单位面积稻田碳吸收强度则表现为增长趋势;碳生产效率呈现缓慢增加的趋势,碳经济效率随着年份递进增加幅度较大,年均增长率为9.9%,碳生态效率则表现为稳定且较低,保持在2.4kg C·kg-1CE左右.表明湖南近几年双季稻生产系统碳综合效率提高缓慢,降低肥料和农药的投入量,提高利用效率是提高双季稻生产系统碳效率的关键.     

不同抗性猕猴桃品种感染溃疡病前后几种保护酶活性变化

研究了不同猕猴桃品种展叶孕蕾期自然感染溃疡病菌前后一年生枝条、叶片内过氧化物酶(POD)、多酚氧化酶(PPO)、超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、苯丙氨酸解氨酶(PAL)酶活性的变化情况.结果表明:感染溃疡病菌前后,抗病品种和感病品种枝条及成叶中防御酶系活性变化规律存在一定的差异.品种未受溃疡病菌感染时,一年生枝条、叶片中的POD、PPO、SOD、CAT酶活性抗病品种的酶活性均低于感病品种.品种自然感染溃疡病菌后,一年生枝条、叶片POD、PPO、SOD、CAT、PAL酶活性均升高,且酶活性的增加幅度是抗病品种中酶活提高倍数高于感病品种.而且这种增量在不同保护酶类以及不同组织部位是有差异的.     

醋酸钙复盐对微生物生长的影响

应用四醋酸钙对黑曲霉、黄曲霉、米曲霉、链格孢霉等霉菌和大肠杆菌、枯草杆菌、绿脓杆菌、金黄色葡萄球菌等细菌生长影响作了研究,并将四醋酸钙与醋酸钙、苯甲酸钠对上述微生物的生长抑制作用加以比较。0.4％四醋酸钙对上述细菌的生长有明显的抑制作用。对上述霉菌生长亦有一定的抑制作用。当其与苯甲酸钠协同使用时,能更明显地抑制微生物生长。实验结果表明四醋酸钙很可能是一种安全性高、成本低的新型食品防腐剂。     

Phylogenetic analysis of Nosema antheraeae (Microsporidia) isolated from Chinese oak silkworm, Antheraea pernyi

The microsporidian Nosema antheraeae is a pathogen that infects the Chinese oak silkworm, Antheraea pernyi. We sequenced the complete small subunit (SSU) rRNA gene and the internal transcribed spacer (ITS) of N. antheraeae, and compared the SSU rRNA sequences in other microsporidia. The results indicated that Nosema species, including N. antheraeae, formed two distinct clades, consistent with previous observations. Furthermore, N. antheraeae is clustered with N. bombycis with high bootstrap support. The organization of the rRNA gene of N. antheraeae is LSU-ITS1-SSU-ITS2-5S, also following a pattern similar to the Nosema type species, N. bombycis. Thus, N. antheraeae is a Nosema species and has a close relationship to N. bombycis.