Development of a novel system to study hepatitis delta virus genome replication.
Hepatitis delta virus (HDV) genome replication requires the virus-encoded small delta protein (deltaAg). During replication, nucleotide sequence changes accumulate on the HDV RNA, leading to the translation of deltaAg species that are nonfunctional or even inhibitory. A replication system was devised where all deltaAg was conditionally provided from a separate and unchanging source. A line of human embryonic kidney cells was stably transfected with a single copy of cDNA encoding small deltaAg, with expression under tetracycline (TET) control. Next, HDV genome replication was initiated in these cells by transfection with a mutated RNA unable to express deltaAg. Thus, replication of this RNA was under control of the TET-inducible deltaAg. In the absence of TET, there was sufficient deltaAg to allow a low level of HDV replication that could be maintained for at least 1 year. When TET was added, both deltaAg and genomic RNA increased dramatically within 2 days. With clones of such cells, designated 293-HDV, the burst of HDV RNA replication interfered with cell cycling. Within 2 days, there was a fivefold enhancement of G1/G0 cells relative to both S and G2/M cells, and by 6 days, there was extensive cell detachment and death. These findings and those of other studies that are under way demonstrate the potential applications of this experimental system.
Chang
Jinhong
J
Fox Chase Cancer Center, 333 Cottman Avenue, Philadelphia, Pennsylvania 19111-2497, USA.
Gudima
Severin O
SO
Tarn
Chi
C
Nie
Xingcao
X
Taylor
John M
JM
D002460:Cell Line; D016679:Genome, Viral; D003698:Hepatitis Delta Virus / Q000187:drug effects / Q000235:genetics* / Q000502:physiology; D006801:Humans; D007668:Kidney; D012367:RNA, Viral / Q000187:drug effects / Q000235:genetics*; D013752:Tetracycline / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects; D014162:Transfection; D014779:Virus Replication
D012367:RNA, Viral; D013752:Tetracycline