Structural requirements of the higher order RNA kissing element in the enteroviral 3'UTR.
The origin of replication ( oriR ) involved in the initiation of (-) strand enterovirus RNA synthesis is a quasi-globular multi-domain RNA structure which is maintained by a tertiary kissing interaction. The kissing interaction is formed by base pairing of complementary sequences within the predominant hairpin-loop structures of the enteroviral 3' untranslated region. In this report, we have fully characterised the kissing interaction. Site-directed mutations which affected the different base pairs involved in the kissing interaction were generated in an infectious coxsackie B3 virus cDNA clone. The kissing interaction appeared to consist of 6 bp. Distortion of the interaction by mispairing of each of the base pairs involved in this higher order RNA structure resulted in either temperature sensitive or lethal phenotypes. The nucleotide constitution of the base which gaps the major groove of the kissing domain was not relevant for virus growth. The reciprocal exchange of the complete sequence involved in the kissing resulted in a mutant virus with wild type virus growth characteristics arguing that the base pair constitution is of less importance for the initiation of (-) strand RNA synthesis than the existence of the tertiary structure itself.
Wang
J
J
University of Nijmegen, Department of Medical Microbiology, PO Box 9101, 6500 HB Nijmegen, The Netherlands.
Bakkers
J M
JM
Galama
J M
JM
Bruins Slot
H J
HJ
Pilipenko
E V
EV
Agol
V I
VI
Melchers
W J
WJ
D020413:3' Untranslated Regions; D000818:Animals; D001483:Base Sequence; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D003198:Computer Simulation; D018076:DNA, Complementary / Q000235:genetics; D029822:Enterovirus B, Human / Q000235:genetics* / Q000254:growth & development; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D012367:RNA, Viral / Q000737:chemistry* / Q000235:genetics; D016415:Sequence Alignment; D014709:Vero Cells; D014779:Virus Replication / Q000235:genetics
D020413:3' Untranslated Regions; D018076:DNA, Complementary; D012367:RNA, Viral