A host-specific function is required for ligation of a wide variety of ribozyme-processed RNAs.
Hepatitis delta virus (HDV) replicates its circular RNA genome via a rolling circle mechanism. During this process, cis-acting ribozymes cleave adjacent upstream sequences and thereby resolve replication intermediates to unit-length RNA. The subsequent ligation of these 5'OH and 2',3'-cyclic phosphate termini to form circular RNA is an essential step in the life cycle of the virus. Here we present evidence for the involvement of a host activity in the ligation of HDV RNA. We used both HDV and hammerhead ribozymes to generate a panel of HDV and non-HDV RNA substrates that bear 5' hydroxyl and 2', 3'- cyclic phosphate termini. We found that ligation of these substrates occurred in host cells, but not in vitro or in Escherichia coli. The host-specific ligation activity was capable of joining RNA in both bimolecular and intramolecular reactions and functioned in a sequence-independent manner. We conclude that mammalian cells contain a default pathway that efficiently circularizes ribozyme processed RNAs. This pathway could be exploited in the delivery of stable antisense and decoy RNA to the nucleus.
Reid
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Department of Molecular Biology, Raymond and Beverly Sackler Research Foundation Laboratory, Tufts University School of Medicine, Boston, MA 02111, USA.
Lazinski
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D001483:Base Sequence; D015152:Blotting, Northern; D002460:Cell Line; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D003698:Hepatitis Delta Virus / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D011118:Polynucleotide Ligases / Q000378:metabolism; D012313:RNA / Q000378:metabolism*; D012322:RNA Precursors / Q000378:metabolism; D016337:RNA, Catalytic / Q000378:metabolism*; D012367:RNA, Viral / Q000378:metabolism; D014764:Viral Proteins
D012322:RNA Precursors; D016337:RNA, Catalytic; D012367:RNA, Viral; D014764:Viral Proteins; D012313:RNA; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases; D011118:Polynucleotide Ligases