REGULATORY ELEMENTS OF AVIAN SARCOMA VIRUS LTRS

禽肉瘤病毒 LTRS 的调控元件

• 批准号: 2376858
• 负责人: RAMAREDDY V GUNTAKA
• 金额: $ 14.18万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-05-01 至 1999-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2376858
• 关键词: RNase protection assay; Rous sarcoma virus; antibody; chemical binding; chickens; complementary DNA; fibroblasts; gel mobility shift assay; gene deletion mutation; gene expression; genetic regulation; genetic regulatory element; genome; host organism interaction; molecular cloning; nucleic acid repetitive sequence; phosphorylation; point mutation; polymerase chain reaction; posttranscriptional RNA processing; transcription factor; transfection; virus genetics

The overall objective of this research project is to understand the host transcription factors involved in the determination of species-specific and tissue-specific expression of Rous sarcoma virus genome. The long terminal repeats (LTR) of the virus contain powerful cis-acting elements to which host factors bind and activate transcription. For some time we have been studying in detail the molecular reactions between host factors and cis- sequences in the promoter-enhancer region (U3) of the RSV lTR. We, and others, have found that at least three well-defined cis elements are required for maximal expression of LTR-linked genes and three different factors interact with these motifs. We have also succeeded in isolating three different yet related cDNA clones encoding polypeptides that specifically bind to distinct sequence motifs. We have shown that one of these factors, mainly present in the avian fibroblasts, and in muscle tissue, binds to a specific site, E3/E3-1, localized at nucleotides -151 to -171. The importance of this cis-acting motif for viral LTR function has not ben examined. The aims of the current study are; (i) to define the role of the cis-acting sequence E3/E3-1 (-151 to -171) in LTR function and characterize the protein binding to this sequence as it appears that this factor is present only in avian fibroblasts, the target tissue for efficient expression of viral genome; (ii) to determine the functional domains of the cloned factors and their cognate cis-acting sequences; (iii) to establish the role of the cloned cDNA products in RSV gene expression. In-frame deletions and point mutations will be made in the cDNAs to identify the various DNA binding domains of the factors. We will also address the issue of whether phosphorylation and homo- or heterodimerization of various factors are required for their interaction, using antibodies raised against the purified factors. These studies will provide further insights into the regulation of viral gene expression and the contribution of host factors in determining tissue tropism and species- specific expression of viral genes. Preliminary results indicated that the GT-rich sequence of the U5 region of the lTR contributes to transcription termination and/or the 3' end processing of the RNA. We intend to continue these studies with an objective of identifying precisely the nucleotides involved in the 3' end maturation of the mRNA, which will be determined by S1 protection assays, oligonucleotide-directed mutagenesis and by studies using polymerase chain reaction.

该研究项目的总体目的是了解主机 确定物种特异性和的转录因子 Rous肉瘤病毒基因组的组织特异性表达。 长期 病毒的重复（LTR）包含强大的顺式作用元素 宿主因子结合并激活转录。 一段时间以来 详细研究宿主因子与顺式之间的分子反应 RSV LTR的启动子增强剂区域（U3）中的序列。 我们， 其他，发现至少三个定义明确的顺式元素是 LTR连接基因和三种不同的最大表达所必需的 因素与这些主题相互作用。 我们也成功地隔离了 三个不同但相关的cDNA克隆编码多肽 特别结合不同的序列基序。 我们已经证明了其中一个 这些因素，主要存在于鸟类成纤维细胞和肌肉中 组织与特定位点E3/e3-1结合，位于核苷酸-151至 -171。 此顺式作用基序对病毒LTR功能的重要性具有 没有Ben检查。 当前研究的目的是： （i）定义 顺式作用序列E3/E3-1（-151至-171）在LTR函数和 表征蛋白质与该序列的结合，因为这似乎 因子仅存在于鸟类成纤维细胞中， 病毒基因组的有效表达； （ii）确定功能 克隆因子的域及其同源顺式作用序列； （iii） 建立克隆的cDNA产物在RSV基因表达中的作用。 将在cDNA中进行框内删除和点突变 确定因子的各种DNA结合域。 我们也会 解决磷酸化和同型或同型的问题 它们相互作用需要各种因素的异二聚化， 使用针对纯化因子提出的抗体。 这些研究会 对病毒基因表达的调节和 宿主因素在确定组织的乳房和物种 - 病毒基因的特定表达。 初步结果表明 LTR的U5区域的富含GT序列有助于转录 RNA的终止和/或3'端处理。 我们打算继续 这些研究目的是精确识别核苷酸 参与mRNA的3'末端成熟，这将由 S1保护测定，寡核苷酸指导的诱变和研究 使用聚合酶链反应。

项目成果

The GT-rich sequence in the U5 region of Rous sarcoma virus long terminal repeat is required for transcription termination and 3' processing.

劳斯肉瘤病毒长末端重复序列的 U5 区中富含 GT 的序列是转录终止和 3 加工所必需的。

• 发表时间: 1997
• 期刊: Folia biologica.
• 作者: Cleavinger,PJ;Kandala,JC;Guntaka,RV
• 通讯作者: Guntaka,RV

Cloning of a novel Y-box homology protein (chkYB-1HP) cDNA lacking the cold-shock domain.

克隆缺乏冷休克结构域的新型 Y-box 同源蛋白 (chkYB-1HP) cDNA。

• DOI: 10.1016/s0167-4781(97)00141-3
• 发表时间: 1998
• 期刊: Biochimica et biophysica acta
• 作者: Nambiar,A;Kandala,JC;Svoboda,J;Guntaka,RV
• 通讯作者: Guntaka,RV

Host- and tissue-specific factors that interact with the enhancer sequences of Rous sarcoma virus LTR.

与劳斯肉瘤病毒 LTR 增强子序列相互作用的宿主和组织特异性因子。

• 发表时间: 1994
• 期刊: Folia biologica
• 影响因子: 0.6
• 作者: Guntaka,RV;Kandala,JC;Shin,BA;Nambiar,A;Svoboda,J;Cleavinger,P
• 通讯作者: Cleavinger,P

Cloning of Rous sarcoma virus enhancer factor genes. II. RSV-EF-II, abundantly expressed in fibroblasts and muscle tissue, binds to an octamer sequence, 5'-GTACCACC-3', in the noncoding strand of RSV enhancer.

劳斯肉瘤病毒增强因子基因的克隆。

• DOI: 10.1006/viro.1996.0404
• 发表时间: 1996
• 期刊: Virology.
• 作者: Cleavinger,PJ;Shin,BA;Kandala,JC;Nambiar,A;Swamynathan,SK;Guntaka,RV
• 通讯作者: Guntaka,RV

Targeted disruption of one allele of the Y-box protein gene, Chk-YB-1b, in DT40 cells results in major defects in cell cycle.

DT40 细胞中 Y-box 蛋白基因 Chk-YB-1b 的一个等位基因的靶向破坏会导致细胞周期的重大缺陷。

• DOI: 10.1016/s0006-291x(02)00875-6
• 发表时间: 2002
• 期刊: Biochemical and biophysical research communications
• 影响因子: 3.1
• 作者: Swamynathan,ShivalingappaK;Varma,BalwantkumarR;Weber,KarlT;Guntaka,RamareddyV
• 通讯作者: Guntaka,RamareddyV