Molecular mechanism by which gamma-herpesviruses evade autophagy

γ-疱疹病毒逃避自噬的分子机制

• 批准号: 7449112
• 负责人: Sangita Sinha
• 金额: $ 23.55万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-15 至 2009-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7449112
• 关键词: Acquired Immunodeficiency Syndrome; Affinity; Amino Acid Sequence; Apoptosis; Apoptotic; Autophagocytosis; BH3 Domain; Binding; Biochemical; Biological Assay; Cell Death; Cell physiology; Cells; Complex; Cultured Cells; Data; Defense Mechanisms; Down-Regulation; Epithelial; Epitopes; Future; Goals; Grant; Herpesviridae; Herpesviridae Infections; Herpesvirus Type 3; Homologous Gene; Host Defense Mechanism; Human; Human Herpesvirus 4; Human Herpesvirus 8; Immunocompromised Host; Individual; Infection; Infectious Mononucleosis; Kaposi Sarcoma; Lymphoid; Malignant Neoplasms; Measurement; Mediating; Membrane; Modeling; Molecular; Murine herpesvirus 68; Mutagenesis; Organelles; Pathogenicity; Patients; Pharmaceutical Preparations; Proteins; Proto-Oncogene Proteins c-bcl-2; Public Health; Purpose; Reagent; Research; Role; Simplexvirus; Site; Specificity; Structure; Surface; Techniques; Testing; Therapeutic; Tissues; Transplant Recipients; Vesicle; Viral; Viral Pathogenesis; Virus; base; design; in vivo; inhibitor/antagonist; mimetics; mutant; novel therapeutics; pathogen; prevent; pro-apoptotic protein; protein aggregate; scaffold; small molecule; three dimensional structure; tumor

DESCRIPTION (provided by applicant): The 3-herpesviridae includes widespread, important human pathogens like Epstein-Barr virus which causes infectious mononucleosis and malignant tumors of lymphoid and epithelial tissues and Kaposi Sarcoma virus, which causes Kaposi sarcoma tumors, especially among immunocompromised individuals such as AIDS patients and transplant recipients. 3-Herpesviruses encode homologs of the anti-apoptotic, cellular Bcl-2 protein (c-Bcl-2), presumably to facilitate viral propagation and oncogenicity by blocking cell death. A key mechanism by which anti-apoptotic Bcl-2 homologs prevent apoptosis is by sequestering pro-apoptotic Bcl-2 homologs. Although viral Bcl-2 (v-Bcl-2) and c-Bcl-2 homologs share very low sequence identity, they have very similar three-dimensional structures. Previous structural, biochemical and mutagenic analyses showed that a hydrophobic groove on the surface of anti-apoptotic Bcl-2 homologs, including most v-Bcl-2 homologs, binds the amphipathic BH3 helical domains of pro-apoptotic Bcl-2 homologs. Despite this shared general mechanism for binding BH3 domains, the different Bcl-2 homologs show unique specificity toward various BH3 domains, arising from differences in their amino acid sequences. 3-herpesviruses have been shown to also inhibit autophagy, a vital cellular process by which organelles, large protein aggregates, stable proteins, as well as intracellular pathogens are enclosed in double-membrane vesicles and targeted for lysosomal degradation. Autophagy is normally modulated by the interaction of c-Bcl-2 homologs with a key autophagy effector protein, Beclin 1, and the down-regulation of autophagy by 3-herpesviruses is mediated by the interaction of v-Bcl-2 homologs with Beclin 1. Recently, Beclin 1 was also shown to contain a BH3 domain that constitutes the primary determinant of binding to Bcl-XL and other c-Bcl-2 homologs. Our preliminary data indicates that additional regions of Beclin 1 contiguous with the BH3 domain are also involved in interaction with Bcl-2 homologs. Further, the various c- and v-Bcl-2 homologs bind to overlapping, but not identical, sites on Beclin 1. The goal of this grant is to characterize the differences and also to verify the similarities, in the interaction of Beclin 1 with v-Bcl-2 and c-Bcl-2 homologs. Interactions will first be quantified by biochemical measurements of binding affinity. Structure determination and analyses will provide details of each interaction, identifying key specificity determinants, which will be confirmed by quantifying binding affinity of selected mutants. Finally, ABT737, a peptido-mimetic that inhibits binding of certain c-Bcl-2 homologs and BH3 domains, will be tested for its ability to disrupt the interaction of v-Bcl-2 homologs and Beclin 1. Thus, this research will provide a better understanding of the molecular mechanisms of herpesvirus pathogenicity, the cellular process of autophagy, the interaction between herpesviral and host proteins, and information on how to selectively modify these interactions either by mutagenesis or by small molecule inhibitors, both for the purposes of further research as well as for designing potential therapeutics to treat this medically important group of pathogens. PUBLIC HEALTH RELEVANCE: Important 3-Herpesviruses human pathogens include EBV, which causes infectious mononucleosis and malignant tumors of lymphoid and epithelial tissues, and KSHV, which causes Kaposi sarcoma tumors, especially among immuno-compromised individuals such as AIDS patients and transplant recipients. These viruses have evolved to down-regulate autophagy, a host defense mechanism known to degrade numerous pathogens. This application aims to investigate atomic details of the mechanism by which 3-herpesviruses inhibit autophagy, providing a better understanding of 3-herpesviral pathogenicity.

描述（由申请人提供）：3-疱疹性病毒包括淋巴样和上皮组织的感染性单核细胞和恶性肿瘤的广泛的，例如爱泼斯坦 - 巴尔病毒，以及在受到免疫疗法的患者中受到kaposi sarcomamem plymors and and and and sarcoma的肿瘤病毒，并受到kaposi sarcomagom promant的疗效。 3-疱疹病毒编码抗凋亡，细胞BCL-2蛋白（C-BCL-2）的同源物，大概是通过阻断细胞死亡来促进病毒传播和致癌性。抗凋亡BCL-2同源物预防凋亡的一种关键机制是隔离促凋亡的Bcl-2同源物。尽管病毒BCL-2（V-BCL-2）和C-BCL-2同源物具有非常低的序列身份，但它们具有非常相似的三维结构。以前的结构，生化和诱变分析表明，包括大多数V-BCl-2同源物在内的抗凋亡BCL-2同源物表面上的疏水凹槽结合了Amphipatic BH3螺旋螺旋结构域的促凋亡BCl-2同源物。尽管这种结合BH3结构域的共享一般机制，但不同的Bcl-2同源物显示出对各种BH3结构域的独特特异性，这是由于其氨基酸序列的差异而引起的。 3-疱疹病毒也已显示可抑制自噬，这是一种重要的细胞过程，通过该过程，细胞器，大蛋白质聚集体，稳定蛋白质以及细胞内病原体被封闭在双膜囊泡中，并靶向溶酶体去分解。自噬通常是通过C-BCl-2同源物与关键自噬效应蛋白Beclin 1的相互作用进行调节的，以及3-赫性病毒对自噬对自噬的下调，介导了V-BCl-2同源物与Beclin 1与Beclin 1的相互作用。同源物。我们的初步数据表明，Beclin 1的其他区域与BH3结构域的连续性也参与了与Bcl-2同源物的相互作用。此外，各种C-和V-BCl-2同源物与Beclin 1上的重叠但不相同的位点结合。该赠款的目的是表征差异并验证Beclin 1与V-BCL-2和C-BCL-2同源物的相互作用。相互作用将首先通过结合亲和力的生化测量来量化。结构确定和分析将提供每种相互作用的详细信息，并确定关键特异性决定因素，这将通过量化所选突变体的结合亲和力来确认。 Finally, ABT737, a peptido-mimetic that inhibits binding of certain c-Bcl-2 homologs and BH3 domains, will be tested for its ability to disrupt the interaction of v-Bcl-2 homologs and Beclin 1. Thus, this research will provide a better understanding of the molecular mechanisms of herpesvirus pathogenicity, the cellular process of autophagy, the interaction between herpesviral and host蛋白质以及有关如何通过诱变或小分子抑制剂进行选择性修改这些相互作用的信息，既出于进一步的研究，又是为了设计潜在的治疗剂来治疗这种重要的病原体。 公共卫生相关性：重要的3疱疹病毒人类病原体包括EBV，它会导致感染性单核细胞增多症和淋巴样和上皮组织的恶性肿瘤，以及KSHV，这会导致Kaposi肉瘤肿瘤，尤其是在诸如AIDS患者和移植者等免疫功能低下的个体中。这些病毒已经演变为下调自噬，这是一种已知降解许多病原体的宿主防御机制。该应用程序旨在研究3-疱疹病毒抑制自噬的机制的原子细节，从而更好地理解了对3 hepesviral的致病性。