Flajolet M, Tiollais P, Buendia M A, Fourel G
Unité de Recombinaison et Expression Génétique, INSERM U163, Institut Pasteur, 75724 Paris Cedex 15, France.
J Virol. 1998 Jul;72(7):6175-80. doi: 10.1128/JVI.72.7.6175-6180.1998.
Direct activation of the N-myc2 oncogene by insertion of woodchuck hepatitis virus (WHV) DNA is a major oncogenic step in woodchuck hepatocarcinogenesis. We previously reported that WHV enhancer II (We2), which controls expression of the core/pregenome RNA, can also activate the N-myc2 promoter in hepatoma cell lines. To better define the integrated WHV regulatory sequences responsible for N-myc2 promoter activation in woodchuck liver tumors, we analyzed the structure and enhancer activity of a single viral integrant found at the win locus in tumor 2260T1 and mapping approximately 175 kb 3' of N-myc2. This viral insert was made of 11 concatemerized WHV fragments, 5 of which overlapped with We2 sequences and 1 with WHV sequence homologous to that of hepatitis B virus enhancer I (We1). In transient transfection assays in hepatoma-derived cells, the We2 activator was found to be fully effective only when inserted in close proximity to the N-myc2 promoter whereas the We1 element by itself was apparently devoid of activity. In contrast, the 2260T1 viral insert exhibited a potent enhancer capacity that depended both on multimerized We2 and on We1 sequences. In a survey of different woodchuck hepatomas, both elements were commonly found within integrated viral sequences involved in long-range N-myc2 activation.
通过插入土拨鼠肝炎病毒（WHV）DNA直接激活N-myc2癌基因是土拨鼠肝癌发生过程中的一个主要致癌步骤。我们先前报道，控制核心/前基因组RNA表达的WHV增强子II（We2），也能在肝癌细胞系中激活N-myc2启动子。为了更好地确定在土拨鼠肝肿瘤中负责激活N-myc2启动子的整合WHV调控序列，我们分析了在肿瘤2260T1的win位点发现的一个单一病毒整合体的结构和增强子活性，该整合体位于N-myc2下游约175 kb处。这个病毒插入片段由11个串联的WHV片段组成，其中5个与We2序列重叠，1个与乙肝病毒增强子I（We1）的WHV序列同源。在肝癌衍生细胞的瞬时转染试验中，发现只有当We2激活剂插入到靠近N-myc2启动子的位置时才完全有效，而We1元件本身显然没有活性。相比之下，2260T1病毒插入片段表现出强大的增强子能力，这既依赖于多聚化的We2，也依赖于We1序列。在对不同土拨鼠肝癌的调查中，这两个元件通常都存在于参与N-myc2远距离激活的整合病毒序列中。
