Hadlock K G, Quasney M W, Lutter L C
Cellular and Molecular Biology Program, University of Michigan, Ann Arbor 48109-0010.
J Biol Chem. 1987 Nov 15;262(32):15527-37.
Tumor antigen (T-antigen) of simian virus 40 (SV40) has been shown to have a number of regulatory roles in both viral replication and early viral transcription. However, the nature of its role on late viral transcription remains unclear. We have analyzed for the presence of T-antigen on SV40 late viral transcription complexes which exhibit RNA polymerase II extension activity in vitro. Nuclear extract or glycerol gradient-isolated transcription complexes were treated with either polyclonal or monoclonal antibodies, and the amount of extension activity that could be immunoprecipitated was determined. Anti-T antibody derived from hamster ascites as well as the anti-T monoclonal antibodies PAb 102, 109, 416, and 419 all precipitated 12-29% of viral transcription complex activity. Immunoprecipitation resulted in significant enrichment of transcription complex activity relative to bulk minichromosomes, indicating a preferential association of T-antigen with the late viral transcription complex. This is the first direct demonstration of the presence of T-antigen on the SV40 late transcription complex. Furthermore, the immunoprecipitated transcription complexes exhibited a salt dependence of their in vitro extension activity which was distinct from that of the total complex population, indicating that T-antigen is present on a specific subclass of transcription complexes.
猴病毒40（SV40）的肿瘤抗原（T抗原）已被证明在病毒复制和早期病毒转录中具有多种调节作用。然而，其在晚期病毒转录中的作用性质仍不清楚。我们分析了在体外表现出RNA聚合酶II延伸活性的SV40晚期病毒转录复合物上是否存在T抗原。用多克隆或单克隆抗体处理核提取物或甘油梯度分离的转录复合物，并测定可被免疫沉淀的延伸活性量。源自仓鼠腹水的抗T抗体以及抗T单克隆抗体PAb 102、109、416和419均沉淀了12 - 29%的病毒转录复合物活性。相对于大量微型染色体，免疫沉淀导致转录复合物活性显著富集，表明T抗原与晚期病毒转录复合物存在优先关联。这是首次直接证明SV40晚期转录复合物上存在T抗原。此外，免疫沉淀的转录复合物在体外延伸活性上表现出盐依赖性，这与总复合物群体不同，表明T抗原存在于转录复合物的一个特定亚类上。
