Three intergenic regions of coronavirus mouse hepatitis virus strain A59 genome RNA contain a common nucleotide sequence that is homologous to the 3' end of the viral mRNA leader sequence

Abstract

cDNA clones that represent various portions of the coronavirus mouse hepatitis virus strain A59 genome RNA have been constructed. cDNAs were synthesized by transcription of genome RNA by using either oligo(dT)12.18 or random oligomers of calf thymus DNA as primers. These cDNAs were converted into double-stranded DNA and cloned into pBR322 by standard techniques. The resulting cloned viral DNA fragments were mapped to viral genes by hybridization with Northern blots of intracellular RNA from mouse hepatitis virus strain A59-infected cells. These cDNA clones map in six of the seven viral genes. Clone g344, 1.8 kilobases, is the largest and encompasses gene 5 (which encodes a nonstructural protein) and gene 6 (which encodes the El viral glycoprotein) as well as the intergenic regions preceding genes 5, 6, and 7. Sequencing of parts of this cloned DNA show that these three intergenic regions contain a common 11-nucleotide sequence. This sequence shares homology with the 3' end of the viral mRNA leader sequence. Thus, this common intergenic sequence may contain a binding site for a leader RNA that hybridizes to negative-strand viral RNA at the beginning of each gene to prime mRNA synthesis. The different degrees of homology between the leader and its putative binding site may influence the differential rates of transcription of the various viral mRNAs.

Comparative sequence analysis of the distal one-third of the genomes of a systemic and an enteric ferret coronavirus

“…The genomic organization observed for each virus was consistent with that of a typical coronavirus. Each intact gene was preceded, a few nucleotides upstream its 5 end, by the recognized consensus transcription-regulating sequence (TRS), 5 -CTAAAC-3 , of coronaviruses (Budzilowicz et al, 1985;Horsburgh et al, 1992;Kapke and Brian, 1986;Snijder et al, 2003;Spaan et al, 1988). In both viruses, the TRS for the E and 7b genes each exhibited a one-base mismatch at the 5 end, appearing as 5 -GTAAAC-3 for the former, and 5 -TTAAAC-3 for the latter.…”

Recombination and Coronavirus Defective Interfering RNAs

“…In this scheme, leader sequences are made, probably in excess, and are, by a process of discontinuous, nonprocessive synthesis, translocated (possibly in a complex with the polymerase) to distant sites on the minusstrand template. It was postulated that the priming was directed by base-pairing between the UCUAAAC element within the free leader and complementary promoter sequences on the minus-strand template (58), or by protein-RNA (40,46) or protein-protein interactions between proteins that recognize these elements (45, 59). Since leader-containing transcripts that might function as free leaders are longer than the leader itself (i.e., Ͼ80 nt) (34) and crossover sites map within or even upstream of the UCUAAAC promoter, the action of a 3Ј = 5Ј exonuclease was postulated for the trimming of the primer before continuation of RNA synthesis (46,51).…”

Quantification of individual subgenomic mRNA species during replication of the coronavirus transmissible gastroenteritis virus

“…Previous work on MHV (Budzilowicz et al, 1985;Shieh et al, 1987) had proposed that the extent of base pairing between the TASs and the 3' end of the leader RNA controlled subgenomic mRNA abundance and provided an explanation of why MHV mRNA 7 was the most abundant subgenomic mRNA in MHV-infected cells. We compared the amounts of the TGEV mRNA species, produced throughout the replication cycle, with the degree of base pairing between the TASs and the 3' end of the leader RNA sequence to determine if there was any correlation with respect to the amounts of the mRNA species produced.…”