Analysis for selective ubiquitination of host immune proteins by Bovine Herpes virus type4 proteins

牛疱疹病毒 4 型蛋白对宿主免疫蛋白的选择性泛素化分析

• 批准号: 16580246
• 负责人: TANAKA Yoshikazu
• 金额: $ 2.37万
• 依托单位: Nippon Veterinary & Animal Science University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2004
• 资助国家: 日本
• 起止时间: 2004 至 2005
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-16580246/
• 关键词: Ubiquitin; Proteolysis; Evasion of immune-system; Persistent infection; ウイルス

Bovine herpesvirus type 4 (BHV-4) is classified as a gammaherpesvirus including of Kaposi's sarcoma-associated herpesvirus (KSHV) on the basis of its genomic structure. BHV-4 strains have been isolated from animals with a variety of clinical signs, such as conjunctivitis, pneumonia, metritis, skin lesions, ulcerative mammillitis, enteritis, and tumors of the urinary bladder and rumen. BHV-4 has two kinds of proteins (Bo4 and Bo5) involved in Plant homeodomain (PHD) that is a motif characteristically defined by seven cysteins and a histidine arranged in a C4HC3 consensus. Recently, PHD domains are related with down-regulation of MHC class I surface expression by increasing the rate of endocytosis and by ubiquitination system to escape from host immune systems.We cloned BHV-4 Bo4 and Bo5 genes from BHV-4 cDNA library and expressed each protein in 293T cells. These proteins were unstable by Western blotting analysis. We, therefore, subcloned the genes into HA-tagged EGFP-fusion vectors. Both N-termini fusion proteins were localized in the nuclear and those of C-termini fusion proteins were localized into the cytoplasm by conforcal microscopy analysis. In addition, N-termini proteins of Bo4 and Bo5 have E3 ubiquitin ligase activities to express autoubiquitination but not C-termini proteins. The experiments of coimmunoprecipitation show that the complex of E2 and E3 proteins was not detected by immunoprecipitation-Western blotting analysis. This means that it is hard to precipitate Bo4 and Bo5 proteins with common anti-HA tag antibody. It is necessary that we may make use of liquid chromatography or ultracentrifugation methods to isolate complex.

牛疱疹病毒4型（BHV-4）根据其基因组结构被分类为包括卡波西肉瘤相关疱疹病毒（KSHV）的γ疱疹病毒。已经从具有多种临床症状的动物中分离出BHV-4菌株，所述临床症状例如结膜炎、肺炎、子宫炎、皮肤损伤、溃疡性乳头炎、肠炎以及膀胱和瘤胃肿瘤。BHV-4有两种蛋白（Bo 4和Bo 5）参与植物同源结构域（PHD），植物同源结构域是由七个半胱氨酸和一个组氨酸以C4 HC 3共有序列排列的特征性基序。本研究从BHV-4 cDNA文库中克隆了BHV-4的Bo 4和Bo 5基因，并在293 T细胞中表达。Western blotting分析表明这些蛋白质不稳定。因此，我们将这些基因亚克隆到HA标记的EGFP融合载体中。通过共聚焦显微镜分析，两种N-末端融合蛋白均定位于细胞核中，而C-末端融合蛋白定位于细胞质中。此外，Bo 4和Bo 5的N-末端蛋白具有E3泛素连接酶活性以表达autoubiquitination，但不表达C-末端蛋白。免疫共沉淀实验表明，免疫共沉淀-Western blotting分析未检测到E2和E3蛋白的复合物。这意味着难以用普通抗HA标签抗体沉淀Bo 4和Bo 5蛋白。有必要采用液相色谱法或超离心法对配合物进行分离。

项目成果

Common anti-apoptotic roles of parkin and alpha-synuclein in human dopaminergic cells.

• 发表时间: 2005
• 期刊: Biochemical and biophysical research communications
• 影响因子: 3.1
• 作者: Y. Machida;T. Chiba;A. Takayanagi;Yoshikazu Tanaka;M. Asanuma;N. Ogawa;A. Koyama;T. Iwatsubo;S. Ito;Poul Hening Jansen;N. Shimizu;Keiji Tanaka;Y. Mizuno;N. Hattori

Changes in octivities of enzymes related to energy metabolism in canine lymphoma cells

犬淋巴瘤细胞能量代谢相关酶活性的变化

• 发表时间: 2005
• 期刊: J Vet Med Sci. (In press)
• 作者: Kitazawa T;De Smet B;Verbeke;K.;Depoortere I.;Peeters T.;三澤尚明;Washizu T
• 通讯作者: Washizu T

Canine epidermal Langerhans cells express a and g but not b chains of high-affinity IgE receptor.

犬表皮朗格汉斯细胞表达高亲和力 IgE 受体的 a 链和 g 链，但不表达 b 链。

• 发表时间: 2005
• 期刊: Veterinary Research Communications (In press)
• 作者: Bungo T;Ueda H;Kitazawa T;Taneike T;Furuse M;Khalifa M et al.;Bonkoabra M
• 通讯作者: Bonkoabra M

Changes in peripheral leukocytes enzymes activity and plasma metabolite concentrations in growing Holstein calves

生长荷斯坦犊牛外周血白细胞酶活性和血浆代谢物浓度的变化

• 发表时间: 2006
• 期刊: Research in Veterinary Science 81/1
• 作者: Hasegawa D;et al.;Nobuhiro Kimura;Toshinori Sako;Toshiro Arai;Haruo Hashimoto;Daisuke Hasegawa;Kei Tamura;Toshiro Arai
• 通讯作者: Toshiro Arai

Glycosylation and Glucose Trimming Inhibitors on Human Parainfluenza Virus Type3.

人类副流感病毒 3 型的糖基化和葡萄糖修饰抑制剂。

• 发表时间: 2006
• 期刊: Antiviral Research (In press)
• 作者: Shiina T.;et al.;Hmeno et al.;Tanaka Y
• 通讯作者: Tanaka Y