Gulliver G A, Herber R L, Liem A, Lambert P F
McArdle Laboratory for Cancer Research, University of Wisconsin Medical School, Madison 53706, USA.
J Virol. 1997 Aug;71(8):5905-14. doi: 10.1128/JVI.71.8.5905-5914.1997.
High-risk human papillomavirus type 16 (HPV-16) and HPV-18 are associated with the majority of human cervical carcinomas, and two viral genes, HPV E6 and E7, are commonly found to be expressed in these cancers. The presence of HPV-16 E7 is sufficient to induce epidermal hyperplasia and epithelial tumors in transgenic mice. In this study, we have performed experiments in transgenic mice to determine which domains of E7 contribute to these in vivo properties. The human keratin 14 promoter was used to direct expression of mutant E7 genes to stratified squamous epithelia in mice. The E7 mutants chosen had either an in-frame deletion in the conserved region 2 (CR2) domain, which is required for binding of the retinoblastoma tumor suppressor protein (pRb) and pRb-like proteins, or an in-frame deletion in the E7 CR1 domain. The CR1 domain contributes to cellular transformation at a level other than pRb binding. Four lines of animals transgenic for an HPV-16 E7 harboring a CR1 deletion and five lines harboring a CR2 deletion were generated and were observed for overt and histological phenotypes. A detailed time course analysis was performed to monitor acute effects of wild-type versus mutant E7 on the epidermis, a site of high-level expression. In the transgenic mice with the wild-type E7 gene, age-dependent expression of HPV-16 E7 correlated with the severity of epidermal hyperplasia. Similar age-dependent patterns of expression of the mutant E7 genes failed to result in any phenotypes. In addition, the transgenic mice with a mutant E7 gene did not develop tumors. These experiments indicate that binding and inactivation of pRb and pRb-like proteins through the CR2 domain of E7 are necessary for induction of epidermal hyperplasia and carcinogenesis in mouse skin and also suggest a role for the CR1 domain in the induction of these phenotypes through as-yet-uncharacterized mechanisms.
高危型人乳头瘤病毒16型（HPV - 16）和HPV - 18与大多数人类宫颈癌相关，并且在这些癌症中通常发现两种病毒基因HPV E6和E7会表达。HPV - 16 E7的存在足以在转基因小鼠中诱导表皮增生和上皮肿瘤。在本研究中，我们在转基因小鼠中进行了实验，以确定E7的哪些结构域促成了这些体内特性。人类角蛋白14启动子用于将突变E7基因的表达导向小鼠的复层鳞状上皮。所选择的E7突变体要么在保守区域2（CR2）结构域有框内缺失，该结构域是视网膜母细胞瘤肿瘤抑制蛋白（pRb）和pRb样蛋白结合所必需的，要么在E7 CR1结构域有框内缺失。CR1结构域在除pRb结合之外的水平上促成细胞转化。产生了四株携带CR1缺失的HPV - 16 E7转基因动物品系和五株携带CR2缺失的品系，并观察其明显的和组织学表型。进行了详细的时间进程分析，以监测野生型与突变型E7对表皮（高水平表达的部位）的急性影响。在具有野生型E7基因的转基因小鼠中，HPV - 16 E7的年龄依赖性表达与表皮增生的严重程度相关。突变型E7基因类似的年龄依赖性表达模式未能导致任何表型。此外，具有突变型E7基因的转基因小鼠未发生肿瘤。这些实验表明，通过E7的CR2结构域与pRb和pRb样蛋白结合并使其失活，对于在小鼠皮肤中诱导表皮增生和致癌作用是必需的，并且还表明CR1结构域通过尚未明确的机制在诱导这些表型中发挥作用。
