Coupled transcription-translation of silkworm cytoplasmic polyhedrosis virus injected into oocytes of the frog, Xenopus laevis.
The cytoplasmic polyhedrosis virus of the silkworm, Bombyx mori, contains a ten-segmented, double-stranded RNA genome and five species of polypeptides, V1 (Mr 146,000), V2 (Mr 140,000), V3 (Mr 128,000)), V4 (Mr 62,000), and V5 (Mr 32,000). The virus contains an RNA-dependent RNA polymerase that transcribes the duplex genome RNA to form mRNA either in the infected animals or under appropriate conditions in vitro. We co-microinjected the virus, [alpha-32P]GTP and actinomycin D into oocytes of the frog, Xenopus laevis, and found that at least eight species of mRNA were formed in the oocytes. Virus-injected oocytes were labeled with [35S]methionine and cell extracts were treated with rabbit anti-cytoplasmic polyhedrosis virus immunoglobulin G. Analysis of the immunoprecipitates by sodium dodecyl sulfate polyacrylamide gel electrophoresis revealed that V1, V2, and V3 were produced in the virus-injected oocytes whereas V4 and V5 were not. By injecting the separated double-stranded genome segments immediately after heat-denaturation into oocytes, it was found that V1 was coded for by segment 1 with a chain length of 4.2 kilobase pairs, V2 by segment 2 or 3, whose chain lengths are both 4.0 kilobase pairs, and V3 by segment 4 with a chain length of 3.2 kilobase pairs. These results demonstrate that the Xenopus oocyte is a very useful system for the coupled transcription-translation of double-stranded RNA viruses.
Ikegami
S
S
Department of Applied Biochemistry, Hiroshima University.
Sasaki
S
S
Higaki
T
T
Itoh
N
N
Shinagawa
N
N
D000818:Animals; D012831:Bombyx / Q000382:microbiology*; D003593:Cytoplasm / Q000382:microbiology; D005260:Female; D005814:Genes, Viral; D007304:Insect Viruses / Q000235:genetics*; D008970:Molecular Weight; D009865:Oocytes / Q000378:metabolism*; D014176:Protein Biosynthesis; D012330:RNA, Double-Stranded / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D014158:Transcription, Genetic; D014982:Xenopus laevis
D012330:RNA, Double-Stranded; D012333:RNA, Messenger