A novel cis-acting element facilitates minus-strand DNA synthesis during reverse transcription of the hepatitis B virus genome.
Hepadnaviruses replicate through reverse transcription of an RNA pregenome, resulting in a relaxed circular DNA genome. The first 3 or 4 nucleotides (nt) of minus-strand DNA are synthesized by the use of a bulge in a stem-loop structure near the 5' end of the pregenome as a template. This primer is then transferred to a complementary UUCA motif, termed an acceptor, within DR1* near the 3' end of the viral pregenome via 4-nt homology, and it resumes minus-strand DNA synthesis: this process is termed minus-strand transfer or primer translocation. Aside from the sequence identity of the donor and acceptor, little is known about the sequence elements contributing to minus-strand transfer. Here we report a novel cis-acting element, termed the beta5 region (28 nt in length), located 20 nt upstream of DR1*, that facilitates minus-strand DNA synthesis. The deletion or inversion of the sequence including the beta5 region diminished minus-strand DNA synthesis initiated at DR1*. Furthermore, the insertion of the beta5 region into its own position in a mutant in which the sequences including the beta5 region were replaced restored minus-strand DNA synthesis at DR1*. We speculate that the beta5 region facilitates minus-strand transfer, possibly by bringing the acceptor site in proximity to the donor site via base pairing or by interacting with protein factors involved in this process.
Shin
Myeong-Kyun
MK
Department of Biochemistry, Yonsei University, 134 Shinchondong, Seodaemungu, Seoul 120-749, Korea.
Lee
Jehan
J
Ryu
Wang-Shick
WS
D045744:Cell Line, Tumor; D004261:DNA Replication; D004279:DNA, Viral / Q000096:biosynthesis*; D004742:Enhancer Elements, Genetic / Q000235:genetics*; D017353:Gene Deletion; D015967:Gene Expression Regulation, Viral; D016679:Genome, Viral; D006515:Hepatitis B virus / Q000235:genetics*; D006801:Humans; D010957:Plasmids; D014158:Transcription, Genetic
D004279:DNA, Viral