NOVEL CELL CYCLE REGULATION BY ADENOVIRUS E1B 55K

腺病毒E1B 55-kDa （55K）蛋白在病毒生长和病毒介导的细胞转化过程中发挥作用。在病毒生长过程中，55K蛋白与E4orf6蛋白结合，调控mRNA的生物发生。在转化过程中，55K蛋白结合肿瘤抑制因子p53并阻断p53应答基因的转录，包括那些促进细胞凋亡和诱导G1生长停滞的基因。然而，大多数感染55k突变病毒的细胞不会发生凋亡，不会在G1期停滞，也不会无法复制突变病毒DNA。因此，55K在病毒生长和转化中的功能之间的联系尚不清楚。这项工作将阐明这种联系，作为理解腺病毒靶向细胞生长控制病毒生长机制的更大目标的一部分。因为55K突变病毒只在S期早期感染的HeLa细胞中生长，而不是在G1期，55K蛋白克服了细胞周期对病毒生长的限制。这个限制和克服这个限制的55K函数将由三个特定的目标决定。首先，通过比较相关g1限制性腺病毒突变体的生长和对病毒生长允许和限制的感染细胞之间mRNA转运的控制，将评估腺病毒必须篡夺mRNA控制细胞周期独立生长的假设。其次，允许型和限制性细胞的异核体将被用来阐明这种限制的细胞基础，并确定限制性表型的优势。第三，鉴定55K/E4orf6复合体靶向的细胞周期无关病毒生长的细胞蛋白，并确定这些蛋白的细胞周期调控。这项研究将确定55K蛋白如何破坏病毒生长的细胞周期控制。这可能代表了55K蛋白在裂解生长和病毒转化之间的第一个联系。这些研究将增加我们对病毒发病机制和病毒肿瘤发生机制以及细胞生长控制的基本机制的认识。此外，本研究将鉴定在S期细胞中生长受限的腺病毒突变体。这种具有复制能力的病毒可用作溶瘤剂来治疗快速生长的人类肿瘤。

The adenovirus E1B 55-kDa (55K) protein functions during virus growth and virus-mediated cellular transformation. During virus growth, the 55K protein binds the E4orf6 protein and regulates mRNA biogenesis. During transformation, the 55K protein binds the tumor suppressor p53 and blocks transcription from p53-responsive genes including those that promote apoptosis and induce G1 growth arrest. However, most cells infected with the 55K-mutant virus do not undergo apoptosis, do not arrest in G1, and do not fail to replicate mutant viral DNA. Consequently, the link between 55K function in virus growth and transformation remains unclear. This work will elucidate this link as part of the larger goal of understanding the means by which adenovirus targets mechanisms of cellular growth control for virus growth. Because 55K-mutant viruses grow only in HeLa cells that were infected in early S phase but not in G1, the 55K protein overcomes a restriction imposed on virus growth by the cell cycle. This restriction and the 55K function that overcomes this restriction will be determined by three specific aims. First, the hypothesis that adenovirus must usurp the control of mRNA for cell cycle-independent growth will be evaluated by comparing the growth of related G1-restricted adenovirus mutants and the control of mRNA transport among infected cells that are permissive and restrictive for virus growth. Second, heterokaryons of permissive and restrictive cells will be used to elucidate the cellular basis for this restriction and determine the dominance of the restrictive phenotype. Third, cellular proteins targeted by 55K/E4orf6 complex for cell cycle-independent virus growth will be identified and the cell cycle- regulation of these proteins determined. This study will determine how the 55K protein subverts cell cycle controls for virus growth. This may represent the first link between the roll of the 55K protein in lytic growth and viral transformation. These studies will increase our understanding of the mechanisms of viral pathogenesis and viral oncogenesis as well as fundamental mechanisms of cellular growth control. Furthermore, this study will identify adenovirus mutants that are restricted for growth in S phase cells. Such replication competent viruses can be used as oncolytic agents to treat rapidly growing human tumors.