カイコ核多角体病ウイルス遺伝子の構造と機能の解析

家蚕核型多角体病毒基因结构与功能分析

• 批准号: 05660388
• 负责人: KOBAYASHI Michihiro
• 金额: $ 1.34万
• 依托单位: Nagoya University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1993
• 资助国家: 日本
• 起止时间: 1993 至 1994
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-05660388/
• 关键词: Silkworm; Nuclear polyhedrosis Virus; DNA polymerase; Gene; Expression

A gene encoding a putative DNA polymerase (pol) of Bombyx mori nuclear polyhedrosis virus (BmSNPV) was cloned and sequenced. The gene included an open reading frame (ORF) encoding a polypeptide of 988 amino acids with a predicted molecular mass of 114.65 kDa. The deduced amino acid sequence of the BmSNPV pol ORF showed an overall identity of 96 and 45% to those of the Autographa californica NPV (AcMNPV) pol ORF and the Lymantria dispar NPV pol ORF,respectively, and contained sequences conserved in a variety of eukaryotic and viral replicative DNA polymerases. The BmSNPV pol lacked a canonical TATAA element but contained a G+C-rich sequence in the transcriptional initiation region. Analyzes by Northern blot hybridization, RNase protection assay, primer extension, and 3' and 5' RACE (rapid amplification of cDNA ends) showed that at least seven different transcripts of approximately 3.1 kb that shared a common 3' end were expressed from the BmSNPV pol. The expression of these transcripts … More from BmSNPV pol was regulated differentially during virus infection. Transcription of five of the seven species initiated in the close vicinity of and within the motif 5'-GCGTGCT-3'. One transcript placed its initiation site within the motif 5'-AGAGCGT-3' and the remaining one within the motif 5'-GGCGGTGG-3'. The motifs 5'-GCGTGCT-3' and 5'-AGAGCGT-3' have been identified in pol and other genes of AcMNPV as conserved sequences containing transcriptional initiation sites, whereas the motif 5'-GGCGGTGG-3', which is arranged as a direct repeat in BmSNPV pol but not in AcMNPV pol, has not been defined as the sequence responsible for transcriptional initiation sites. The BmSNPV pol transcripts were detectable at 2 hr postinfection (p.i.) , peaked at 10 hr p.i., and declined to a low level by 18 hr p.i. The expression of BmSNPV pol was not inhibited but delayd dramatically by the protein synthesis inhibitor cycloheximide upon treatment of infected cells, whereas aphidicolin, an inhibitor of DNA polymerase, inhibited BmSNPV pol transcription. These results suggest a complicated and unique mechanism for the regulation of BmSNPV pol expression. Less

对家蚕核型多角体病毒(BmSNPV)DNA聚合酶基因进行了克隆和序列分析。该基因包括一个开放阅读框，编码一个由988个氨基酸组成的多肽，预测的分子质量为114.65 kDa。BmSNPV polORF的氨基酸序列与银纹夜蛾核型多角体病毒(Autograph a calfornica NPV，AcMNPV)polORF和毒蛾核型多角体病毒(Lymantria Disdisar NPV)polORF的氨基酸序列同源性分别为96%和45%，并含有多种真核生物和病毒复制DNA聚合酶中保守的序列。BmSNPV pol1缺乏典型的TATAA元件，但在转录起始区含有富含G+C的序列。经Northern杂交、RNase保护分析、引物延伸和3‘和5’RACE(快速扩增cDNAEnds)分析表明，BmSNPV Poll至少表达了7个约3.1kb的共同3‘端的不同转录本。这些转录本…的表达在病毒感染过程中，更多的BmSNPV Poll受到不同程度的调控。7个物种中有5个物种的转录起始于5‘-GCGTGCT-3’基序附近和内部。一个转录本将其起始位置放在5‘-AGAGCGT-3’基序中，其余一个转录本放在5‘-GGCGGTGG-3’基序中。5‘-GCGTGCT-3’和5‘-AGAGCGT-3’基序在Pol和AcMNPV的其他基因中已被确定为含有转录起始点的保守序列，而在BmSNPV pol1中以直接重复的形式排列的5‘-GGCGGTGG-3’基序尚未被定义为负责转录起始点的序列。BmSNPV Poll转录本在感染后2小时可检测到，10小时达到高峰，18小时下降到较低水平。蛋白质合成抑制剂放线菌亚胺处理感染细胞后，BmSNPV pol1基因的表达不受抑制，但显著延迟，而DNA聚合酶抑制剂aphidiclin则能抑制BmSNPV pol1基因的转录。这些结果提示了BmSNPV pol1基因表达调控的复杂而独特的机制。较少

项目成果

Sotoshiro, H.and Kobayashi, M.: "Identification of viral structural polypeptides in the midgut and feces of the silkworm, Bombyx mori, infected with Bombyx densovirus type 2." J.Invertebr.Pathol.(in press).

Sotoshiro, H. 和 Kobayashi, M.：“感染 2 型家蚕浓核病毒的家蚕中肠和粪便中病毒结构多肽的鉴定。”

Chaeychomsri, S., Ikeda, M.and Kobayashi, M.: "Nucleotide sequence and transcriptional analysis of the DNA polymerase gene of Bombyx mori nuclear polyhedrosis virus." Virology. 206. 435-447 (1995)

Chaeychomsri, S.、Ikeda, M. 和 Kobayashi, M.：“家蚕核多角体病毒 DNA 聚合酶基因的核苷酸序列和转录分析。”

Chaeychomsri, S., Ikeda, M., Kobayashi, M.: "Expression of viral DNA polymerase gene in the larvae of the silkworm, Bombyx mori (Lepidoptera : Bombycidae), during the infection with nuclear polyhedrosis virus." Appl.Entomol.Zool.(in press).

Chaeychomsri, S.、Ikeda, M.、Kobayashi, M.：“在感染核型多角体病毒期间，家蚕（鳞翅目：家蚕科）幼虫中病毒 DNA 聚合酶基因的表达。”

Kobayashi,M.and Caeychomsri,S.: "Thermal inhibition of viral disease in the silkworm,Bombyx mori" Indian Journal of Sericulture. 32. 1-7 (1993)

Kobayashi,M. 和 Caeychomsri,S.：“家蚕病毒性疾病的热抑制，Bombyx mori”《印度蚕业杂志》。

Kobayashi, M.and Chaeychomsri, S.: "Thermal inhibition of viral disease in the silkworm" Bombyx mori.Indian J.Sericult.32. 1-7 (1993)

Kobayashi, M. 和 Chaeychomsri, S.：“蚕病毒性疾病的热抑制”Bombyx mori.Indian J.Sericult.32。