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Regulation of human cytomegalovirus strain AD169 gene expression

By Gavin William Grahame Wilkinson


Human cytomegalovirus (HCMV) strain AD169 encodes a single abundant 1.95kb immediate early (IE) mRNA and a single abundant 2.7kb early RNA. The major IE gene (0.756-0.745 map units) was shown in nuclease protection experiments to encode a spliced molecule of 1,736 nucleotides (excluding the poly(A) tail) consisting of four exons of 121, 88, 185 and 1,342 nucleotides. Three introns of 827, 114 and 170 nucleotides were located near the 5' end of the gene. The structural analysis of the major IE gene enabled the amino acid sequence of the major IE polypeptide to be deduced from the DNA sequence. The major early gene, which is contained in both copies of the HCMV long repeat, was found not to be spliced. A translation product of the 2.7kb early RNA has yet to be identified. Reporter genes were used in transient DNA transfection experiments to monitor expression from HCMV and other viral promoters. HCMV infections trans-activated expression from the transfected SV40 early, Rous Sarcoma virus, HSV-1 thymidine kinase (TK), the HCMV major IE and the HCMV major early promoters. Expression from both the HCMV IE and the HSV-1 TK promoters was stimulated much more gradually by HCMV than by HSV-1 infections. Experiments performed using u.v.-irradiated virus and inhibitors of HCMV replication indicated that the transfected IE promoter was stimulated primarily by a de novo synthesised HCMV-encoded gene product(s). When the concentration of the plasmids IEPlcatIEterm and AccHincat transfected into cells was lowered sufficiently, HCMV infection was observed to repress expression from the transfected IE promoter. A sequence in the HCMV major IE gene between -299 and + 69 apparently contains a cis-acting signal which responds to an HCMV-induced repressor. Competitive co-transfection experiments indicated that an HCMV-induced repressor of IE transcription interacts with at least three distinct regions within the IE promoter

Topics: QR355
OAI identifier: oai:wrap.warwick.ac.uk:2540

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