Base-pairing promotes leader selection to prime in vitro influenza genome transcription.
The requirements for alignment of capped leader sequences along the viral genome during influenza transcription initiation (cap-snatching) have long been an enigma. In this study, competition experiments using an in vitro transcription assay revealed that influenza virus transcriptase prefers leader sequences with base complementarity to the 3'-ultimate residues of the viral template, 10 or 11 nt from the 5' cap. Internal priming at the 3'-penultimate residue, as well as prime-and-realign was observed. The nucleotide identity immediately 5' of the base-pairing residues also affected cap donor usage. Application to the in vitro system of RNA molecules with increased base complementarity to the viral RNA template showed stronger reduction of globin RNA leader initiated influenza transcription compared to those with a single base-pairing possibility. Altogether the results indicated an optimal cap donor consensus sequence of (7m)G-(N)(7-8)-(A/U/G)-(A/U)-AGC-3'.
Laboratory of Virology, Wageningen University, Droevendaalsesteeg 1, 6708 PB Wageningen, The Netherlands.
Christina
C
Geerts-Dimitriadou
Mark P
MP
Zwart
Rob
R
Goldbach
Richard
R
Kormelink
D020121:5' Untranslated Regions / Q000235:genetics* / Q000502:physiology; D017794:Alfalfa mosaic virus / Q000235:genetics / Q000378:metabolism; D000818:Animals; D020029:Base Pairing / Q000235:genetics* / Q000502:physiology; D001483:Base Sequence; D016679:Genome, Viral / Q000235:genetics / Q000502:physiology; D006801:Humans; D053118:Influenza A Virus, H1N1 Subtype / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D012315:RNA Caps / Q000235:genetics* / Q000502:physiology; D012367:RNA, Viral / Q000235:genetics / Q000502:physiology; D011817:Rabbits; D013698:Templates, Genetic; D014158:Transcription, Genetic
D020121:5' Untranslated Regions; D012315:RNA Caps; D012367:RNA, Viral