Jariwalla R J, Aurelian L, Ts'o P O
Proc Natl Acad Sci U S A. 1983 Oct;80(19):5902-6. doi: 10.1073/pnas.80.19.5902.
Diploid Syrian hamster embryo (SHE) cells were passaged after transfection with recombinant plasmids containing herpes simplex virus type 2 (HSV-2) DNA inserts Bgl II focus-forming fragment N, Bgl II transforming fragment C, and EcoRI/HindIII fragment AE. Cultures transfected with salmon DNA or with 0.1-5.0 micrograms of Bgl II fragment N reached crisis and senesced. Those transfected with 0.1-0.5 micrograms of Bgl II fragment C or its left-hand 64% subclone EcoRI/HindIII fragment AE escaped senescence and formed continuous lines. At early passages, these lines as well as isolated clones grew in 2% serum but formed small (less than or equal to 0.1 mm) colonies in 0.3% agarose and were nontumorigenic. Serial passaging of Bgl II fragment C-induced cultures and isolated clones resulted in the appearance of large (greater than 0.25 mm) colonies in agarose followed by tumorigenicity. This behavior was not exhibited by the EcoRI/HindIII fragment AE-induced cultures that remained nontumorigenic after 53 passages. DNA from normal SHE cells exhibited homology to Bgl II fragment C but, under relatively stringent conditions, DNAs from transformed and tumor-derived lines exhibited discrete hybridizing bands comigrating with authentic viral fragments. These results indicate that neoplastic transformation of normal diploid SHE cells by HSV-2 DNA fragments involves at least two distinct steps--i.e., immortalization and conversion to tumorigenicity. EcoRI/HindIII fragment AE representing the left 64% of Bgl II fragment C is sufficient to induce immortalization. However, DNA sequences from both left-hand 64% and right-hand 36% subfragments of Bgl II fragment C are required for tumorigenic transformation.
将含有单纯疱疹病毒2型（HSV - 2）DNA插入片段Bgl II成灶性形成片段N、Bgl II转化片段C以及EcoRI/HindIII片段AE的重组质粒转染二倍体叙利亚仓鼠胚胎（SHE）细胞后进行传代培养。用鲑鱼DNA或0.1 - 5.0微克Bgl II片段N转染的培养物进入危机期并衰老。用0.1 - 0.5微克Bgl II片段C或其左侧64%的亚克隆EcoRI/HindIII片段AE转染的细胞逃脱了衰老并形成了连续细胞系。在早期传代时，这些细胞系以及分离的克隆在2%血清中生长，但在0.3%琼脂糖中形成小（≤0.1毫米）菌落，且无致瘤性。对Bgl II片段C诱导的培养物和分离的克隆进行连续传代，导致在琼脂糖中出现大（＞0.25毫米）菌落，随后具有致瘤性。EcoRI/HindIII片段AE诱导的培养物在传代53次后仍无致瘤性，未表现出这种行为。正常SHE细胞的DNA与Bgl II片段C具有同源性，但在相对严格的条件下，来自转化细胞系和肿瘤衍生细胞系的DNA显示出与真实病毒片段共迁移的离散杂交带。这些结果表明，HSV - 2 DNA片段对正常二倍体SHE细胞的肿瘤转化涉及至少两个不同步骤，即永生化和转化为致瘤性。代表Bgl II片段C左侧64%的EcoRI/HindIII片段AE足以诱导永生化。然而，Bgl II片段C的左侧64%和右侧36%亚片段的DNA序列对于致瘤性转化都是必需的。
