Temporal expression profile of late gene expression factor 4 from Bombyx mori nucleopolyhedrovirus.
Temporal expression profile of lef4, the gene encoding late gene expression factor 4 (LEF4) from the baculovirus, Bombyx mori nucleopolyhedrovirus (BmNPV), has been analysed. lef4 behaved like an early gene and the transcripts were detectable from 6 h post infection (hpi) which reached maximal levels by 18-24 hpi, and declined considerably at later times. The LEF4 open reading frame was bacterially expressed as a glutathione S-transferase (GST) fusion protein which was solubilized from the inclusion bodies and purified by adsorption to the affinity matrix, GST-Sepharose. Using polyclonal antibodies raised against the bacterially expressed protein, the temporal profile of LEF4 synthesis in BmNPV-infected BmN cells was analysed. The LEF4 protein levels were also higher at 24 hpi compared to 12 or 36 hpi, correlating with the RNA patterns. The protein was predominantly localized to the nucleus of the infected BmN cell and only a small portion was present in the cytosolic fraction. Preliminary studies with antisense lef4 expression revealed substantial reduction in expression from the viral polyhedrin promoter without significantly affecting the viral DNA replication.
Sehrawat
Seema
S
Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India.
Gopinathan
Karumathil P
KP
D000818:Animals; D012831:Bombyx / Q000166:cytology / Q000821:virology*; D002460:Cell Line; D003001:Cloning, Molecular; D016373:DNA, Antisense / Q000235:genetics; D004279:DNA, Viral / Q000737:chemistry / Q000235:genetics; D019084:Fluorescent Antibody Technique, Indirect; D015967:Gene Expression Regulation, Viral; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D017924:Nucleopolyhedroviruses / Q000235:genetics* / Q000254:growth & development; D011401:Promoter Regions, Genetic / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA; D013997:Time Factors; D014158:Transcription, Genetic; D014162:Transfection; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism
D016373:DNA, Antisense; D004279:DNA, Viral; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; C471155:late gene expression factor 4, Bombyx mori nucleopolyhedrovirus; D005982:Glutathione Transferase