A short region in the genome of hepatitis B virus is critical for maintenance of high transcript levels.
The majority of hepatitis B virus (HBV) transcripts are not normally spliced during the viral life cycle, but several splice donor and acceptor sites are conserved on HBV transcripts. In particular, the genome region between nt 450 and 500 of the HBV genome appears to be rich in such sequences. In this study we deleted a short 30-nt sequence between a conserved splice donor site at HBV genome position 462 and a splice acceptor site at position 491, thus deleting the surface/polymerase open reading frames by 10 amino acid residues. At the transcriptional level, this deletion led to >99% reduction of the 2.1-kb class of subgenomic transcripts in transfected cells. Nuclear run-on experiments revealed that the transcription rate of the deleted 2.1-kb transcript is unchanged when compared with the wildtype, suggesting a posttranscriptional mechanism for the downregulation of the deleted transcript. In addition, experiments with a replication-competent HBV mutant containing the 30-nt deletion showed that the corresponding 10-amino-acid sequence within the reverse transcriptase domain of the polymerase protein appeared to be nonessential.
zu Putlitz
J
J
Molecular Hepatology Laboratory, Massachusetts General Hospital Cancer Center and Harvard Medical School, Boston, Massachusetts, 02129, USA.
Tong
S
S
Wands
J R
JR
D000595:Amino Acid Sequence; D001483:Base Sequence; D002460:Cell Line; D004261:DNA Replication; D004279:DNA, Viral / Q000096:biosynthesis; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D006513:Hepatitis B e Antigens / Q000235:genetics*; D006515:Hepatitis B virus / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D011061:Poly A; D016133:Polymerase Chain Reaction; D012326:RNA Splicing; D016415:Sequence Alignment; D014158:Transcription, Genetic