Evolution of the 5′-end of genomic rna of murine coronaviruses during passages in vitro()

The 5′-ends of the genomic RNA and subgenomic mRNAs of murine coronavirus (MHV) have a stretch of approximately 70 nucleotides of leader sequences. The 3'-region of this leader sequence contains several repeats of a pentanucleotide (UCUAA), whose number varies among different MHV strains. It has been demonstrated that this UCUAA repeat plays crucial roles in the discontinuous transcription of MHV mRNAs. In the present study, we demonstrate that the number of UCUAA repeats in the leader sequence of MHV genome rapidly decreases during serial passages of viruses on susceptible cells. The downward evolution of the number of UCUAA repeats was not due to a higher growth rate of the viruses with fewer repeats, but seemed to be due to homologous interference between viruses with different numbers of UCUAA repeat. The ease with which these variant viruses arose suggests the high frequency of the occurrence of this deletion during RNA replication. This finding is in agreement with the proposed discontinuous and nonprocessive mode of coronavirus RNA synthesis. Analysis of the intracellular subgenomic mRNA species of viruses with different numbers of UCUAA repeats and of MHV recombinant viruses suggests that the number of this pentanucleotide repeat at the 3′-end of the leader sequence may regulate the synthesis of certain mRNA species, in agreement with the leader-primed transcription mechanism.