Distinct requirement for two stages of protein-primed initiation of reverse transcription in hepadnaviruses.
Reverse transcription in hepadnaviruses is primed by the viral reverse transcriptase (RT) (protein priming) and requires the specific interaction between the RT and a viral RNA signal termed epsilon, which bears the specific template sequence for protein priming. The product of protein priming is a short oligodeoxynucleotide which represents the 5' end of the viral minus-strand DNA and is covalently attached to the RT. We have now identified truncated RT variants from the duck hepatitis B virus that were fully active in the initial step of protein priming, i.e., the covalent attachment of the first nucleotide to the protein (RT deoxynucleotidylation), but defective in any subsequent DNA polymerization. A short sequence in the RT domain was localized that was dispensable for RT deoxynucleotidylation but essential for the subsequent DNA polymerization. These results have thus revealed two distinct stages of protein priming, i.e., the initial attachment of the first nucleotide to the RT (RT deoxynucleotidylation or initiation of protein priming) and the subsequent DNA synthesis (polymerization) to complete protein priming, with the second step entailing additional RT sequences. Two models are proposed to explain the observed differential sequence requirement for the two distinct stages of the protein priming reaction.
Wang
Xingtai
X
Department of Microbiology, Boston University School of Medicine, Boston, Massachusetts 02118, USA.
Hu
Jianming
J
D020121:5' Untranslated Regions / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D004279:DNA, Viral / Q000378:metabolism; D015611:Hepatitis B Virus, Duck / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides / Q000378:metabolism*; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism; D012194:RNA-Directed DNA Polymerase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic
D020121:5' Untranslated Regions; D004279:DNA, Viral; D009838:Oligodeoxyribonucleotides; D012367:RNA, Viral; D012194:RNA-Directed DNA Polymerase