AMINOACYL-TRNAS IN HIV AND OTHER RETROVIRAL INFECTED CELLS

本项目的主要目标是确定宿主氨酰-tRNA 在人类免疫缺陷病毒中， 病毒（HIV）和其它逆转录病毒被改变（即，低修饰的）从 正常细胞的氨酰转运RNA 初步证据显示 一个实验室提出，HIV和其他逆转录病毒中的这些tRNA 受感染的细胞是低修饰的。 小鼠乳腺肿瘤病毒（MMTV） gag-pro移码信号（序列A AAA AAC，其中C是gag-pro移码信号的位点）。 移码）被用作这些研究的模型。 得出 野生型移码信号和一系列移码突变 作为一种工具，用于确定是否需要tRNA改变MMTV中的 阅读框架是低修饰的。 移帧的级别降低 即使AAC/AAU密码子被编码， 相同天冬酰胺同种受体对其它mRNA具有相似的效率 网站. 低修饰的天冬酰胺tRNA缺乏高度修饰的Q 已知碱基优先读取AAC而不是AAU密码子。 因此 观察到在移码位点的AAC密码子促进核糖体 比AAU密码子更有效的移码表明， 解码移帧信号需要缺少Q基。 我们有 迄今为止，还没有成功地利用体外测定来证明 在核糖体移码中需要低修饰的同种受体 由于测定系统中的内源性tRNA群体。 但我们 已经开发了一种核糖体移码的体内测定， 显微注射技术注入非洲爪蟾卵母细胞， 纯化的低修饰同种受体改变阅读的能力 野生型和突变型MMTV gag-pro移码信号的框。

The major goal of this project is to determine if host aminoacyl-tRNAs required for decoding the frameshift signal in human immunodeficient virus (HIV) and other retroviruses are altered (i.e., hypomodified) from the normal cellular aminoacyl-tRNAs. Preliminary evidence from this laboratory has suggested that these tRNAs in HIV and other retroviral infected cells are hypomodified. The mouse mammary tumor virus (MMTV) gag-pro frameshift signal (sequence A AAA AAC, where C is the site of the frameshift) is being used as a model for these studies. We obtained the wild type frameshift signal and a series of mutations at the frameshift site in MMTV as a tool for determining if the tRNA required to alter the reading frame is hypomodified. The level of frameshifting is reduced dramatically with an AAU mutant even though AAC/AAU codons are decoded by the same asparagine isoacceptor with similar efficiencies at other mRNA sites. Hypomodified asparagine tRNA which lacks the highly modified Q base is known to preferentially read AAC over AAU codons. Thus, the observation that the AAC codon at the frameshift site promotes ribosomal frameshifting more efficiently than the AAU codon suggests that the tRNA lacking Q base is required for decoding the frameshift signal. We have not been successful in utilizing in vitro assays thus far to demonstrate a requirement for hypomodified isoacceptors in ribosomal frameshifting due to the endogenous tRNA population in the assay system. However, we have developed an in vivo assay for ribosomal frameshifting utilizing microinjection techniques into Xenopus oocytes and are presently assaying the ability of purified hypomodified isoacceptors to alter the reading frame of the wild type and mutant MMTV gag-pro frameshift signals.