Formation of a Ribonucleoprotein Complex of Mouse Hepatitis Virus Involving Heterogeneous Nuclear Ribonucleoprotein A1 and Transcription-Regulatory Elements of Viral RNA

Abstract

The heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) specifically binds to two transcription-regulatory elements, i.e., the leader and intergenic sequence, of the negative-strand (template-strand) RNA of mouse hepatitis virus (MHV) and may play a role in viral RNA transcription. Previous studies based on the defective-interfering RNAs of MHV suggested that these two RNA elements may interact with each other during transcription, although they do not have complementary sequences. In this study, we showed by an in vitro reconstitution assay that hnRNP A1 could mediate the formation of an RNP complex involving these two RNA elements. Both the RNA-binding domains and protein-interacting domain of hnRNP A1 contributed to the efficient formation of the RNP complex; however, the presence of the two RNA-binding domains alone, without the protein-interacting domain, also resulted in some RNP formation. Omission of hnRNP A1 in the reconstitution reaction abolished the RNP formation, and mutations of the IG sequences significantly inhibited the RNP formation. These findings suggest that the two cis-acting transcription-regulatory sequences of MHV RNA can interact with each other through the formation of an RNP complex involving a cellular protein hnRNP A1. This RNP complex may participate in MHV RNA transcription.