Stem-loop III in the 5' untranslated region is a cis-acting element in bovine coronavirus defective interfering RNA replication.
Higher-order structures in the 5' untranslated region (UTR) of plus-strand RNA viruses are known in many cases to function as cis-acting elements in RNA translation, replication, or transcription. Here we describe evidence supporting the structure and a cis-acting function in defective interfering (DI) RNA replication of stem-loop III, the third of four predicted higher-order structures mapping within the 210-nucleotide (nt) 5' UTR of the 32-kb bovine coronavirus (BCoV) genome. Stem-loop III maps at nt 97 through 116, has a calculated free energy of -9.1 kcal/mol in the positive strand and -3.0 kcal/mol in the negative strand, and has associated with it beginning at nt 100 an open reading frame (ORF) potentially encoding an 8-amino-acid peptide. Stem-loop III is presumed to function in the positive strand, but its strand of action has not been established. Stem-loop III (i) shows phylogenetic conservation among group 2 coronaviruses and appears to have a homolog in coronavirus groups 1 and 3, (ii) has in all coronaviruses for which sequence is known a closely associated short, AUG-initiated intra-5' UTR ORF, (iii) is supported by enzyme structure-probing evidence in BCoV RNA, (iv) must maintain stem integrity for DI RNA replication in BCoV DI RNA, and (v) shows a positive correlation between maintenance of the short ORF and maximal DI RNA accumulation in BCoV DI RNA. These results indicate that stem-loop III in the BCoV 5' UTR is a cis-acting element for DI RNA replication and that its associated intra-5' UTR ORF may function to enhance replication. It is postulated that these two elements function similarly in the virus genome.
Raman
Sharmila
S
Department of Microbiology, University of Tennessee, College of Veterinary Medicine, Knoxville, Tennessee 37996-0845, USA.
Bouma
Peter
P
Williams
Gwyn D
GD
Brian
David A
DA
D020121:5' Untranslated Regions / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002417:Cattle; D017938:Coronavirus, Bovine / Q000235:genetics* / Q000378:metabolism; D003673:Defective Viruses / Q000235:genetics* / Q000378:metabolism; D004742:Enhancer Elements, Genetic; D016679:Genome, Viral; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D034622:RNA Interference; D012367:RNA, Viral / Q000096:biosynthesis* / Q000235:genetics; D014779:Virus Replication
D020121:5' Untranslated Regions; D012367:RNA, Viral