STRUCTURE AND EXPRESSION OF 2 DEFECTIVE ADENOVIRUS 2-SIMIAN VIRUS 40 HYBRIDS
STRUCTURE AND EXPRESSION OF 2 DEFECTIVE ADENOVIRUS 2-SIMIAN VIRUS 40 HYBRIDS
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DOI：
10.1016/0022-2836(78)90065-7
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发表时间：
1978-01-01
影响因子：
5.6
通讯作者：
SAMBROOK, J
中科院分区：
文献类型：
--
作者：
HASSELL, JA;LUKANIDIN, E;SAMBROOK, J
从称为Ad2 ++ HEY的群体中分离出两种新的缺陷型腺病毒2/SV-40杂交体（Ad2 + D1和Ad2 + D2）。通过限制性内切酶分析、杂交和电镜观察，确定了它们的基因组结构。Ad2 + D1的DNA大小几乎与其辅助腺病毒2的大小相等。它含有插入的SV-40序列3.2x。103个碱基长，代替3.5 × 103个碱基长。腺病毒2型DNA的103个碱基的片段，其定位在距病毒DNA左端0.64 - 0.74个分数基因组长度之间。包含SV-40基因组的整个早期区域的插入物包括位于位置71和10之间的DNA片段：它以编码A基因蛋白质的羧基末端的序列向左定向。Ad2 + D2的DNA比腺病毒2的DNA短约6%。从腺病毒2DNA左端开始的0.76 - 0.96分数基因组长度之间的区段图是不存在的：在其位置插入一段DNA，5.0 × 105。长度为103个碱基，其包含SV-40基因组的所有序列，除了位于图谱位置54 - 63之间的那些序列。与Ad2 + D1相比，Ad2 + D2中的早期SV-40序列从左至右转录。在Ad2 + D1感染[非洲绿色]猴[肾CV-1]细胞后的早期和晚期，细胞质中存在与SV-40 DNA E链早期序列互补的RNA。未检测到病毒L链的转录物。SV-40特异性RNA在Ad2 + D2感染的细胞的细胞质中被发现，仅在感染后的晚期。最丰富的物种是与SV-40 DNA的E链编码序列互补的。L链的转录本也存在，但数量明显较少。SV-40特异性T [肿瘤]抗原相关的蛋白质在感染的细胞中检测到免疫荧光和免疫沉淀与血清从仓鼠荷瘤诱导SV-40。从Ad2 + D1感染的细胞提取物中沉淀的主要多肽以与SV-40 T抗原相同的速率迁移通过十二烷基硫酸钠/聚丙烯酰胺凝胶：其表观MW为96，000。从Ad2 + D2感染的细胞提取物中沉淀出明显更大的多肽（MW = 115，000）。对Ad2 + D1和Ad2 + D2基因组结构的考虑表明，Ad2 + D1可能是由Ad2 ++ HEY杂交池成员与腺病毒2之间的重组产生的，而Ad2 + D2最有可能是由腺病毒2与游离SV-40之间的重组事件产生的。
Two new defective adenovirus 2/SV-40 hybrids (Ad2+Dl and Ad2+D2) were isolated from the population known as Ad2++HEY. The structure of their genomes was determined by analysis with restriction enzymes, hybridization and EM. The DNA of Ad2+D1 is almost equal in size to that of its helper, adenovirus 2. It contains an insertion of SV-40 sequences 3.2 .times. 103 bases long in place of the 3.5 .times. 103 base segment of adenovirus 2 DNA which maps between 0.64-0.74 fractional genome lengths from the left end of the viral DNA. The insertion which encompasses the entire early region of the SV-40 genome, comprises the segment of DNA lying between positions 71 and 10: it is oriented with the sequences coding for the carboxy terminus of the A gene protein to the left. The DNA of Ad2+D2 is shorter than that of adenovirus 2 by approximately 6%. The segment mapping between 0.76-0.96 fractional genome lengths from the left end of adenovirus 2 DNA is absent: in its place is inserted a stretch of DNA, 5.0 .times. 103 bases in length which comprises all the sequences of the SV-40 genome apart from those lying between map positions 54-63. By contrast to Ad2+D1, the early SV-40 sequences in Ad2+D2 are transcribed from left to right. RNA complementary to the early sequences of the E strand of SV-40 DNA is present in the cytoplasm at both early and late times after infection of [African green] monkey [kidney CV-1] cells by Ad2+D1. No transcripts of the viral L strand were detected. SV-40-specific RNA was found in the cytoplasm of cells infected with Ad2+D2, only at late times after infection. The most abundant species was complementary to the coding sequences of the E strand of SV-40 DNA. Transcripts of the L strand were also present, but in significantly lesser quantity. Proteins related to SV-40-specific T [tumor] antigen were detected in infected cells by immunofluorescence and by immunoprecipitation with sera from hamsters bearing tumors induced by SV-40. The major polypeptide precipitated from extracts of cells infected with Ad2+D1 migrated through sodium dodecyl sulfate/polyacrylamide gels at the same rate as SV-40 T antigen: its apparent MW is 96,000. A significantly larger polypeptide (MW = 115,000) is precipitated from extracts of cells infected by Ad2+D2. Consideration of the structures of the genomes of Ad2+D1 and Ad2+D2 indicate that Ad2+D1 may have arisen by recombination between members of the Ad2++HEY hybrid pool and adenovirus 2, while Ad2+D2 was most likely generated by recombination events between adenovirus 2 and free SV-40.