期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：1991
卷号：88
期号：16
页码：7140-7144
DOI：10.1073/pnas.88.16.7140
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：In human cells infected with adenovirus, the virus-associated RNA VAI blocks the activation of the interferon-induced double-stranded-RNA-dependent 68-kDa protein kinase (p68) and maintains normal levels of protein synthesis at late times after infection. VAI antagonizes the kinase activity by binding to p68. The structure of VAI consists of two long, base-paired stems connected by a complex short stem-loop structure. Previous work using a series of adenovirus mutants showed that the structural determinants of the VAI RNA that are essential for function reside in the central complex short stem-loop structure and adjacent base-paired regions (functional domain); the long duplex regions were found to be dispensable for function. To determine whether binding of VAI to p68 correlates with function and whether the structural determinants that are essential for function are also essential for binding, we studied the interaction of wild-type and several mutant VAI RNAs with p68 in whole cells. The p68-VAI complexes from mutant- and wild-type-infected cells were immunoprecipitated by an anti-p68 monoclonal antibody. The mutant RNAs that functioned efficiently in the cells bound to p68 efficiently in the cells, whereas functionally impaired mutants failed to bind to p68, indicating that the binding of the VAI RNA to p68 correlates well with function. In vitro binding assays with immunopurified p68 confirmed these observations. Secondary-structure analysis of several mutant VAI RNAs suggests that the binding does not depend on the long duplex regions but requires all the elements of the functional domain. We propose that the functional domain and the p68-binding domain of the VAI RNA are identical.
