Regulation of NF-kappaB-dependent lymphocyte activation and development by paracaspase

Paracaspase (MALT1), a member of an evolutionarily conserved superfamily of caspase-like proteins, has been shown to bind and colocalize with the protein Bcl10 in vitro and, because of this association, has been suggested to be involved in the CARMA1-Bcl10 pathway of antigen-induced nuclear factor kappaB (NF-kappaB) activation. We demonstrate that primary T and B lymphocytes from paracaspase-deficient mice are defective in antigen-receptor-induced NF-kappaB activation, cytokine production, and proliferation. Paracaspase acts downstream of Bcl10 to induce NF-kappaB activation and is required for the normal development of B cells, indicating that paracaspase provides the missing link between Bcl10 and activation of the IkappaB kinase complex.     

Analysis of junctional diversity during B lymphocyte development

Immunoglobulin rearrangement is central to generating antibody diversity because of heterogeneity generated during recombination by deletion or addition of nucleotides at coding joints by the recombinase machinery. Examination of these junctional modifications revealed that the addition of nongermline-encoded nucleotides was more prevalent in adult versus fetal B cells, thus partially limiting the fetal antibody repertoire. In contrast, deletion of nucleotides occurs equivalently in B cells at different stages of development and at different points in B cell ontogeny. Finally, the bias in murine immunoglobulins for one DH segment reading frame occurs at the DHJH intermediate.     

Regulation of phospholipase C-gamma 1 by profilin and tyrosine phosphorylation.

Epidermal growth factor and platelet-derived growth factor can stimulate the production of the second messenger inositol trisphosphate in responsive cells, but the biochemical pathway for these signaling events has been uncertain because the reactions have not been reconstituted with purified molecules in vitro. A reconstitution is described that requires not only the growth factor, its receptor with tyrosine kinase activity, and the soluble phospholipase C-gamma 1, but also the small soluble actin-binding protein profilin. Profilin binds to the substrate phosphatidylinositol 4,5-bisphosphate and inhibits its hydrolysis by unphosphorylated phospholipase C-gamma 1. Phosphorylation of phospholipase C-gamma 1 by the epidermal growth factor receptor tyrosine kinase overcomes the inhibitory effect of profilin and results in an effective activation of phospholipase C-gamma 1.     

Regulation of JNK by Src during Drosophila development

In Drosophila, the Jun amino-terminal kinase (JNK) homolog Basket (Bsk) is required for epidermal closure. Mutants for Src42A, a Drosophila c-src protooncogene homolog, are described. Src42A functions in epidermal closure during both embryogenesis and metamorphosis. The severity of the epidermal closure defect in the Src42A mutant depended on the amount of Bsk activity, and the amount of Bsk activity depended on the amount of Src42A. Thus, activation of the Bsk pathway is required downstream of Src42A in epidermal closure. This work confirms mammalian studies that demonstrated a physiological link between Src and JNK.     

维生素B1对油菜氧化胁迫的调节作用

［目的］研究VB1对甲基紫精（MV）介导的油菜幼苗氧化胁迫伤害的影响。［方法］以油菜种子为试验材料,用4种不同浓度的MS、MV、VB1及MV＋VB1培养液处理油菜种子,通过测定不同处理下的油菜幼苗鲜重和植物体内SOD、CAT和APX活性以及丙二醛（MDA）和脯氨酸（Pro）的含量,研究VB1对缓解油菜幼苗氧化胁迫作用的影响。［结果］MV可抑制油菜植株的生长,诱导植株体内丙二醛（MDA）和脯氨酸（Pro）含量的增高以及CAT、SOD和APX酶活性的增加;VB1在一定程度上缓解MV介导的油菜幼苗氧化胁迫的伤害,其缓解程度大小与VB1的浓度有关,当VB1的浓度为2.0 mmol/L时,缓解作用最大。［结论］VB1对MV引起的油菜幼苗氧化胁迫有一定的调节作用,其作用大小与MV和VB1的浓度有关。     

月光花素调控下甘薯的生长发育

用天然生长调节物质月光花素处理徐薯１８的研究结果表明，经０．１活性单位月光花素处理，甘薯插枝发根量增加１培多，根系活力明显增强；植株生长快，叶面积增加２０％—６０％，高叶面积时期从１个月延长到２个月，因而有效光合叶面积和光合时间明显增加．合理施用月光花素，可使幼薯分化提早１０天，薯数增加１８％，薯块肥大加速，约可提前２０天成熟，甘薯产量平均可提高１４％，相当于每公顷净增鲜薯４５００ｋｇ，作者在文中还比较了在甘薯生长的不同发育时期，用不同方法施用不同浓度的月光花素，对甘薯生长发育的影响。     

Tuberculin-active carbohydrate that induces inhibition of macrophage migration but not lymphocyte transformation

A tuberculin carbohydrate fraction, GAE, in sensitized animals induced a delayed type of skin reactivity and inhibited the migration of macrophages but failed to stimulate lymphocyte transformation in vitro. Tuberculin protein-containing fractions were active in each test. These results show that in vitro lymphocyte transformation is not necessarily a corollary of delayed type hypersensitivity.     

紫薇花器官发育调控基因LiFUL1的分离与表达分析

紫薇湘韵具有只开花不结实的特性,表现为花药不开裂、花粉败育、胚珠发育异常。紫薇红叶为普通紫薇,可正常开花结实。以紫薇湘韵为试验材料,采用RT-PCR方法,从花芽中分离得到1个FRUITFULL(FUL)同源基因,命名为LiFUL1(GenBank登录号为MN894547)。序列分析结果表明,其cDNA开放阅读框长度为753 bp,编码251个氨基酸,分子质量为28.453 13 ku。序列比对和保守结构域分析结果表明,LiFUL1基因编码蛋白具有典型的MADS-MEF2和K-box结构域,C末端含有一个保守性高的基序euFUL MOTIF,因此,LiFUL1属于MADS家族的FUL/AP1亚家族。进化分析表明,紫薇的LiFUL1基因编码的蛋白氨基酸序列与木本植物蓝桉的FUL/AP1氨基酸序列亲缘关系最近。qRT-PCR分析结果表明,LiFUL1基因在紫薇湘韵花器官分化阶段的花萼分化时期、花瓣与雄蕊分化时期、雄蕊与雌蕊分化时期的表达量显著高于紫薇红叶。另外,利用原核表达系统在大肠埃希菌中成功表达了LiFUL1蛋白,为进一步研究LiFUL1基因的功能奠定了基础。     

Alteration of lymphocyte trafficking by sphingosine-1-phosphate receptor agonists

Blood lymphocyte numbers, essential for the development of efficient immune responses, are maintained by recirculation through secondary lymphoid organs. We show that lymphocyte trafficking is altered by the lysophospholipid sphingosine-1-phosphate (S1P) and by a phosphoryl metabolite of the immunosuppressive agent FTY720. Both species were high-affinity agonists of at least four of the five S1P receptors. These agonists produce lymphopenia in blood and thoracic duct lymph by sequestration of lymphocytes in lymph nodes, but not spleen. S1P receptor agonists induced emptying of lymphoid sinuses by retention of lymphocytes on the abluminal side of sinus-lining endothelium and inhibition of egress into lymph. Inhibition of lymphocyte recirculation by activation of S1P receptors may result in therapeutically useful immunosuppression.     

多个环境因素对嗜水气单胞菌感染团头鲂致病力的影响

目的研究pH对拟态弧菌(Vibrio mimicus)感染相关表型及胞外产物特性的影响。方法测定不同pH (6.0、7.0、8.0、8.5和9.0)下拟态弧菌的增殖、泳动、自聚集和生物膜形成以及胞外产物(extracellular products,ECPs)的蛋白质量浓度、酶活性、溶血性、细胞毒性和毒力等特性。结果pH 6.0~9.0范围内,随着pH值升高,拟态弧菌的增殖先增强后减弱,pH 8.5时增殖最强;自聚集、生物膜形成和ECPs毒力先减弱后增强,pH 8.5时显著减弱(P<0.05);泳动以及ECPs蛋白质量浓度和细胞毒性减弱,pH 6.0时显著强于其他pH (P<0.05);ECPs酶活性和溶血性呈波动性,酶活性在pH 6.0时最强、pH 8.0时最弱,溶血性在pH 8.0时最强、pH 8.5时最弱。转录组分析显示：拟态弧菌pH 8.5较pH 7.0时,与ATP合成相关的糖酵解、TCA循环和氧化磷酸化等过程的基因表达上调,而与ADP合成相关的嘌呤合成基因、与蛋白质合成相关的核糖体亚基合成基因以及与蛋白分泌和膜运输相关的基因表达下调。结论pH对拟态弧菌感染相关表型及ECPs特性具有明显影响,且pH可能是通过调节拟态弧菌能量代谢与膜运输等过程影响其相关生物学特性。     

丙型肝炎病毒NS4B蛋白调控Hep3B细胞非折叠蛋白质反应研究

研究表达的丙型肝炎病毒(HCV)NS4B对Hep3B细胞非折叠蛋白质反应的影响。NS4B重组真核表达质粒pcDNA3.1(-)NS4B通过脂质体转染Hep3B细胞,G418筛选和Western blot鉴定稳定转染细胞;RT-PCR检测稳定转染细胞内XBP1 mRNA剪接,Western blot鉴定ATF6蛋白切割,荧光素酶试验检测稳定转染细胞内GRP78和XBP1启动子活性。G418筛选和Western Blot鉴定证实获得稳定表达NS4B的Hep3B细胞;在该细胞内,XBP1 mRNA剪接、ATF6切割、XBP1和Grp78启动子激活均被检测到。NS4B在Hep3B的稳定表达诱导了非折叠蛋白质反应。     

Regulation of CD8+ T cell development by thymus-specific proteasomes

Proteasomes are responsible for generating peptides presented by the class I major histocompatibility complex (MHC) molecules of the immune system. Here, we report the identification of a previously unrecognized catalytic subunit called beta5t. beta5t is expressed exclusively in cortical thymic epithelial cells, which are responsible for the positive selection of developing thymocytes. Although the chymotrypsin-like activity of proteasomes is considered to be important for the production of peptides with high affinities for MHC class I clefts, incorporation of beta5t into proteasomes in place of beta5 or beta5i selectively reduces this activity. We also found that beta5t-deficient mice displayed defective development of CD8(+) T cells in the thymus. Our results suggest a key role for beta5t in generating the MHC class I-restricted CD8(+) T cell repertoire during thymic selection.     

雄激素受体及共调节因子在睾酮调节绵羊附睾GPX5表达中的作用分析

为探究雄激素受体(Androgen receptor,AR)及共调节因子对绵羊附睾上皮细胞(Epididymal epithelial cells,EECs)谷胱甘肽过氧化物酶5(Glutathione peroxidase 5,GPX5)的调节机制,本研究采用CCK-8检测睾酮对EECs增殖的影响;利用qRT-PCR、免疫荧光法和 Western Blot法分别检测AR、GPX5、甾体激素受体共激活子 1(Steroid receptor coactivator 1,SRC-1)、CREB结合蛋白(cAMP-response element-binding protein,CBP)、p300和核受体共抑制因子2(Nuclear receptor corepressor 2,NCOR2)的mRNA水平和蛋白表达情况,采用双荧光素酶报告基因测定干扰SRC-1或p300后EECs的荧光素酶活性。结果表明:1)与对照组相比,100 nmol/L睾酮组细胞中的GPX5 mRNA和蛋白表达量极显著升高(P<0.01),1 000 nmol/L睾酮组细胞中的GPX5 mRNA和蛋白表达量差异不显著(P>0.05),但分别极显著(P<0.01)和显著(P<0.05)低于100 nmol/L睾酮组;100 nmol/L睾酮组细胞中的AR mRNA和蛋白表达量分别呈极显著(P<0.01)和显著(P<0.05)高于对照组,然而1 000 nmol/L睾酮组细胞中的AR mRNA和蛋白的表达量显著低于对照组(P<0.05);1 000 nmol/L睾酮组细胞中的CBP、p300、SRC-1蛋白表达极显著高于对照组(P<0.01),特别是核内的表达量明显增高。2)与对照组相比,siRNA-AR组细胞中的GPX5 mRNA和蛋白的表达量差异不显著(P>0.05),而SRC-1和p300蛋白表达量极显著升高(P<0.01)。pcDNA3.1-AR组GPX5 mRNA和蛋白表达量较对照组呈极显著(P<0.01)和显著(P<0.05)升高。3)siRNA-SRC-1 和siRNA-p300组细胞中的GPX5 mRNA和蛋白表达量较对照组均显著降低(P<0.05),而AR的蛋白表达量与对照组差异不显著(P>0.05),AR的转录活性显著降低(P<0.05)。综上,睾酮对绵羊EECs GPX5、AR及其共调节因子的表达具有浓度依赖性的调节效应,睾酮通过AR及其共调节因子SRC-1和p300/CBP的协同调节GPX5表达。本研究为探究GPX5在绵羊附睾中的调节机制提供理论依据。     

Inhibition of antigen-induced lymphocyte proliferation by Tat protein from HIV-1

The purified human immunodeficiency virus type-l (HIV-l) Tat protein inhibited lymphocyte proliferation induced by tetanus toxoid or Candida antigens by 66 to 97% at nanomolar concentrations of Tat. In contrast, Tat did not cause a significant reduction of lymphocyte proliferation in response to mitogens such as phytohemagglutinin or pokeweed mitogen. Inhibition was blocked by oxidation of the cysteine-rich region of Tat or by incubation with an antibody to Tat before the assay. A synthetic Tat peptide (residues 1 to 58) also inhibited antigen-stimulated proliferation. Experiments with H9 and U937 cell lines showed that Tat can easily enter both lymphocytes and monocytes. The specific inhibition of antigen-induced lymphocyte proliferation by Tat mimics the effect seen with lymphocytes from HIV-infected individuals and suggests that Tat might directly contribute to the immunosuppression associated with HIV infection.