Krippl B, Ferguson B, Jones N, Rosenberg M, Westphal H
Proc Natl Acad Sci U S A. 1985 Nov;82(22):7480-4. doi: 10.1073/pnas.82.22.7480.
We have modified the E1A gene of human subgroup C adenovirus by introducing deletions in its coding sequence. Various truncated E1A proteins were expressed in Escherichia coli, purified, and microinjected via glass capillaries into Vero cells. We monitored their movement from the cell cytoplasm to the nucleus and their ability to induce expression of H5dl312, an adenovirus E1A deletion mutant. Our results show that the carboxyl terminus of E1A contains sequences essential for rapid and efficient nuclear localization. Essential information for efficient H5dl312 complementation is contained in an internal region, comprising sequences of both exons of the E1A gene. A first exon-encoded region, however, is sufficient to induce low levels of adenovirus gene expression. Information for nuclear localization and for H5dl312 complementation are therefore encoded by distinct domains of the E1A gene. In addition, we determined that the human c-myc product was unable to complement H5dl312.
我们通过在其编码序列中引入缺失来修饰人C亚组腺病毒的E1A基因。各种截短的E1A蛋白在大肠杆菌中表达、纯化，并通过玻璃毛细管显微注射到Vero细胞中。我们监测了它们从细胞质到细胞核的移动以及它们诱导腺病毒E1A缺失突变体H5dl312表达的能力。我们的结果表明，E1A的羧基末端包含快速高效核定位所必需的序列。高效H5dl312互补的关键信息包含在一个内部区域，该区域包括E1A基因两个外显子的序列。然而，第一个外显子编码区域足以诱导低水平的腺病毒基因表达。因此，核定位信息和H5dl312互补信息由E1A基因的不同结构域编码。此外，我们确定人c-myc产物不能互补H5dl312。
