Braspenning J, Marchini A, Albarani V, Levy L, Ciccolini F, Cremonesi C, Ralston R, Gissmann L, Tommasino M
Angewandte Tumorvirologie, Deutsches Krebsforschungszentrum, Heidelberg, Germany.
Oncogene. 1998 Feb 26;16(8):1085-9. doi: 10.1038/sj.onc.1201617.
The conserved region 3 (CR3) of the E7 protein of human papillomaviruses contains two CXXC motifs involved in zinc binding and in the homodimerization of the molecule. Studies have suggested that the intact CXXC motifs in the CR3 of HPV16 and HPV18 E7 are required for the in vitro transforming activity of these proteins. CR3 also contains a low affinity pRb binding site and is involved in the disruption of the E2F/Rb1 complex. E7 is structurally and functionally related to Adenovirus E1A protein, which also has two CXXC motifs in CR3. However, the Ad E1A transforming activity appears to be independent of the presence of such domains. In fact, this viral protein exists in vivo as two different forms of 289 and 243 amino acids. The shorter Ad E1A form (Ad E1A243), where both CXXC motifs are deleted by internal splicing, retains its in vitro transforming activity. We have investigated if the HPV16 E7 CR3 can be functionally replaced by the Ad E1A243 CR3, which lacks both CXXC motifs. A chimeric protein (E7/E1A243) containing the CR1 and CR2 of HPV16 E7 fused to the CR3 of Ad E1A 243 was constructed. The E7/E1A243 while not able to homodimerize in the S. cerevisiae two-hybrid system retains several of the properties of the parental proteins, HPV16 E7 and Ad E1A. It associates with the 'pocket' proteins, induces growth in soft agar of NIH3T3 cells and immortalizes rat embryo fibroblasts. These data suggest that the CXXC motifs in CR3 of E7 do not play a direct role in the transforming properties of this viral protein but probably are important for maintaining the correct protein configuration.
人乳头瘤病毒E7蛋白的保守区域3（CR3）含有两个参与锌结合和分子同二聚化的CXXC基序。研究表明，HPV16和HPV18 E7的CR3中完整的CXXC基序是这些蛋白体外转化活性所必需的。CR3还含有一个低亲和力的pRb结合位点，并参与E2F/Rb1复合物的破坏。E7在结构和功能上与腺病毒E1A蛋白相关，后者在CR3中也有两个CXXC基序。然而，腺病毒E1A的转化活性似乎与这些结构域的存在无关。事实上，这种病毒蛋白在体内以两种不同形式存在，分别为289个和243个氨基酸。较短的腺病毒E1A形式（腺病毒E1A243），其两个CXXC基序均通过内部剪接缺失，但仍保留其体外转化活性。我们研究了HPV16 E7的CR3是否能被缺乏两个CXXC基序的腺病毒E1A243的CR3功能替代。构建了一种嵌合蛋白（E7/E1A243），它包含HPV16 E7的CR1和CR2，并与腺病毒E1A 243的CR3融合。E7/E1A243虽然在酿酒酵母双杂交系统中不能同二聚化，但保留了亲本蛋白HPV16 E7和腺病毒E1A的一些特性。它与“口袋”蛋白结合，诱导NIH3T3细胞在软琼脂中生长，并使大鼠胚胎成纤维细胞永生化。这些数据表明，E7的CR3中的CXXC基序在这种病毒蛋白的转化特性中不发挥直接作用，但可能对维持正确的蛋白质构象很重要。
