MOLECULAR BIOLOGY AND BIOCHEMICAL STRUCTRUE OF ENDOGENOUS PROVIRUSES OF MICE

小鼠内源性原病毒的分子生物学和生化结构

• 批准号: 4688492
• 负责人: A S KHAN
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/4688492
• 关键词: Retroviridae; chromosomes; gene expression; genetic manipulation; genetic mapping; genetic regulation; genome; molecular biology; molecular cloning; murine leukemia virus; nucleic acid sequence; provirus; simian virus 40; virus DNA; virus antigen; virus envelope

We had previously isolated several endogenous MuLV proviruses from BALB/c and AKR/J mouse DNAS. Molecular and biochemical characterization of the cloned endogenous MuLV DNAs indicated that about 50% were related to known MuLV proviruses (Class I). Restriction enzyme and nucleotide sequence analysis indicated that the Class I endogenous MuLV DNAs could be distinguished from known infectious MuLV proviruses due to the presence of a transposon-like 190 bp cellular insert in their LTRs and unique restriction sites. Furthermore, the env sequences associated with the majority of such endogenous MuLV DNAs were similar to those present in recombinant MCF MuLVs. In fact, one cloned endogenous AKR MuLV DNA, designated as A-12, was almost identical in sequence of its 5' env region with leukemogenic MCF-MuLV env genes. We have determined the genomic location of the A-12 provirus to be on chromosome 13 or 18. Unlike other endogenous MuLV DNAs, the A-12 endogenous provirus was present in several inbred as well as wild mouse DNAs. Nucleotide sequence comparison of the LTR, 3' pol and env regions of leukemogenic (MCF-13) and non-leukemogenic (MCF-111A) MuLVs indicated a high degree of base homology in the env region. The LTR of MCF-13 was highly related to the LTR present in xenotropic MuLVs whereas, the LTR associated with MCF-111A was identical except for 1 bp with ecotropic MuLV proviral LTRs. The 3' pol region of MCF-13 contained a 12 bp sequence characteristic of leukemogenic MCF MuLVs. This was absent in MCF-111A MuLV DNA. The leukemogenic potential of in vitro constructed recombinant MuLVs was tested. These viruses contained 5' MCF-13 LTR, gag, pol and env sequences and 3' endogenous MuLV env and LTR sequences. Newborn AKR mice inoculated with such recombinant viruses developed thymic lymphomas in about 3 months. Tissue-specific expression of endogenous MCF-related mRNAs was detected in various AKR/N mouse tissues using P32-labeled MCF env-specific synthetic DNA probe. A 7.2 kb mRNA species was identified in the thymus which could be involved in the generation of recombinant thymotropic MCF viruses.

我们之前已经从 BALB/c 中分离出几种内源性 MuLV 原病毒 和 AKR/J 小鼠 DNAS。 分子和生化特征 克隆的内源 MuLV DNA 表明大约 50% 与已知的 MuLV 原病毒（I 类）。 限制性内切酶和核苷酸序列 分析表明，I 类内源 MuLV DNA 可能是 由于存在以下特征，与已知的传染性 MuLV 原病毒区分开来： 在其 LTR 中插入类似转座子的 190 bp 细胞插入物，并且是独特的 限制位点。 此外，与相关的 env 序列 大多数此类内源性 MuLV DNA 与存在于 重组MCF MuLV。 事实上，一种克隆的内源 AKR MuLV DNA， 指定为 A-12，其 5' env 区域的序列几乎相同 具有致白血病 MCF-MuLV env 基因。 我们已经确定了基因组 A-12 原病毒的位置位于 13 或 18 号染色体上。与其他病毒不同 内源性 MuLV DNA 中，A-12 内源性原病毒存在于多种病毒中 近交系和野生小鼠 DNA。 核苷酸序列比较 白血病 (MCF-13) 和非白血病的 LTR、3' pol 和 env 区域 (MCF-111A) MuLV 表明环境中具有高度的碱基同源性 地区。 MCF-13 的 LTR 与 中存在的 LTR 高度相关 异向性 MuLV，而与 MCF-111A 相关的 LTR 是相同的 除了亲嗜性 MuLV 原病毒 LTR 的 1 bp 外。 3' pol 区域 MCF-13 含有白血病 MCF 的 12 bp 序列特征 MuLV。 MCF-111A MuLV DNA 中不存在这一点。 致癌潜力 测试了体外构建的重组 MuLV。 这些病毒 包含 5' MCF-13 LTR、gag、pol 和 env 序列以及 3' 内源 MuLV env 和 LTR 序列。 接种这种重组体的新生 AKR 小鼠 病毒在大约 3 个月内发展为胸腺淋巴瘤。 组织特异性 在各种 AKR/N 中检测到内源性 MCF 相关 mRNA 的表达 使用 P32 标记的 MCF env 特异性合成 DNA 探针检测小鼠组织。 一个 在胸腺中鉴定出 7.2 kb mRNA 种类，可能参与 重组促胸腺MCF病毒的产生。