Analysis of cis-Acting Sequences Essential for Coronavirus Defective Interfering RNA Replication

Mouse hepatitis virus (MHV) defective interfering (DI) RNA was used to determine the cis-acting sequences required for MHV RNA replication. A 2.2-kb-long cDNA clone of the MHV DI RNA DIssE was used to test the effect of deletions throughout the DI RNA on replication in DI RNA-transfected, MHV-infected cells. Data from a series of deletion mutants demonstrated that about 470 nucleotides at the 5′ terminus, 460 nucleotides at the 3′ terminus, and about 135 nucleotides in an internal position approximately 0.9 kb from the 5′ end of DI RNA were necessary for DI RNA replication. These data suggested that cis-acting sequences which were necessary for MHV RNA replication required not only terminal sequences but also an internal sequence present at about 3.2 kb from the 5′ end of the genome.