Mutations that increase in situ priming also decrease circularization for duck hepatitis B virus.
The process of hepadnavirus reverse transcription involves two template switches during the synthesis of plus-strand DNA. The first involves translocation of the plus-strand primer from its site of generation, the 3' end of minus-strand DNA, to the complementary sequence DR2, located near the 5' end of the minus-strand DNA. Plus strands initiated from DR2 are extended to the 5' end of the minus-strand DNA. At this point, the 3' end of the minus strand becomes the template via the second template switch, a process called circularization. Elongation of circularized plus-strand DNA generates relaxed circular DNA. Although most virions contain relaxed circular DNA, some contain duplex linear DNA. Duplex linear genomes are synthesized when the plus-strand primer is used at the site of its generation, the 3' end of the minus-strand template. This type of synthesis is called in situ priming. Although in situ priming is normally low, in some duck hepatitis B virus mutants this type of priming is elevated. For example, mutations within the 3' end of the minus-strand DNA can lead to increased levels of in situ priming. We report here that these same mutations result in a second defect, a less efficient template switch that circularizes the genome. Although it is not clear how these mutations affect both steps in DNA replication, our findings suggest a commonality in the mechanism of initiation of plus-strand synthesis and the template switch that circularizes the genome.
Loeb
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McArdle Laboratory for Cancer Research, University of Wisconsin Medical School, 1400 University Ave., Madison, WI 53706, USA. loeb@oncology.wisc.edu
Tian
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D002460:Cell Line; D004261:DNA Replication / Q000235:genetics*; D004279:DNA, Viral / Q000096:biosynthesis* / Q000235:genetics; D015611:Hepatitis B Virus, Duck / Q000235:genetics* / Q000502:physiology; D017354:Point Mutation; D014158:Transcription, Genetic; D014162:Transfection; D014779:Virus Replication
D004279:DNA, Viral