Nascent synthesis of leader sequence-containing subgenomic mRNAs in coronavirus genome-length replicative intermediate RNA.
Infection with coronavirus results in the accumulation of genomic-sized mRNA and six to eight subgenomic mRNAs that make up a 3' coterminal nested-set structure. Genome-length negative-strand RNA and subgenomic-length negative-strand RNAs, each of which corresponds to each of the subgenomic mRNAs, also accumulate in infected cells. The present study examined whether the genome-length negative-strand RNA serves as a template for subgenomic mRNA synthesis. Genome-length replicative intermediate (RI) RNA was purified by two-dimensional gel electrophoresis of intracellular RNAs from cells infected with mouse hepatitis virus. RNase A treatment of the purified genome-length RI resulted in the production of the genome-length replicative form RNA, indicating that the genome-length RI included genome-length template RNA. RNase protection assays using the purified genome-length RI and two probes, which corresponded to the 5' 300-nt region of mRNA 6 and to the same region of mRNA 7, showed the presence of nascent leader sequence-containing subgenomic mRNAs in the genome-length RI. These data demonstrated that the genome-length negative-strand RNA serves as a template for subgenomic mRNA synthesis.
Mizutani
T
T
Department of Microbiology and Immunology, The University of Texas Medical Branch at Galveston, Galveston, Texas 77555-1019, USA.
Repass
J F
JF
Makino
S
S
D000818:Animals; D002460:Cell Line; D004587:Electrophoresis, Agar Gel; D015180:Electrophoresis, Gel, Two-Dimensional; D016679:Genome, Viral; D051379:Mice; D006517:Murine hepatitis virus / Q000235:genetics* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000096:biosynthesis*; D012367:RNA, Viral / Q000096:biosynthesis*; D014158:Transcription, Genetic
D012333:RNA, Messenger; D012367:RNA, Viral