Structure of Adenovirus Virus Associated RNA-I

Protein kinase R (PKR) is activated by dsRNA produced during virus replication and plays a major role in the innate immunity response to virus infection. In response, viruses have evolved multiple strategies to evade PKR. Adenovirus virus-associated RNA I (VAI) is a short, non-coding transcript that function as an RNA decoy to sequester PKR in an inactive state. VAI consist of an apical stem-loop, a highly structured central domain, and a terminal stem. Chemical probing and mutagenesis experiments were used to produce a refined secondary structure of VAI and to probe tertiary structure within the central domain. Nucleotides within two loops are protected from SHAPE modification, indicating that they participate in tertiary interactions. Mutations within either loop designed to disrupt a putative pseudoknot result in enhanced modification at both sites. Introduction of compensatory basepairs induces protection, indicating that these loops interact to form a pseudoknot. The residue-level constraints from SHAPE and low-resolution structural information from SAXS measurements were used to develop an atomic model of VAI. The ab initio model of VAI derived from SAXS measurements contains a central bulged region flanked by a short arm and a longer, kinked arm. A putative atomic model of VAI was predicted based on the experimental constraints from SHAPE analysis using a fragment assembly approach. The atomic structure agrees well with the SAXS envelope. This model rationalizes the roles of the three domains of VAI in mediating high affinity PKR binding.