An element in the bovine papillomavirus late 3' untranslated region reduces polyadenylated cytoplasmic RNA levels.
Expression of the two bovine papillomavirus type 1 (BPV-1) late genes, L1 and L2, coding for the two capsid proteins, is limited to terminally differentiated keratinocytes in bovine fibropapillomas. This pattern of expression is determined both by the activity of the late promoter and by the inhibition of late region expression in less well differentiated cells. Inhibition of L1 and L2 mRNA production in nonpermissive cells must occur since the late region potentially could be transcribed from early region promoters. Nuclear runoff analysis of the late region has demonstrated that up to 95% of transcripts which are initiated in the early region in nonpermissive cells terminate within the late region upstream of the late polyadenylation site (C. C. Baker and J. Noe, J. Virol. 63:3529-3534, 1989). However, very few of the primary transcripts which include the late polyadenylation site are processed into mRNA. In this study, we have used expression vectors to characterize an inhibitory element active in nonpermissive cells which is located in the late 3' untranslated region (3'UTR). While the late polyadenylation site is functional in these cells, a 53-bp element in the late 3'UTR reduces levels of polyadenylated cytoplasmic RNA. This element inhibited chloramphenicol acetyltransferase (CAT) expression 6- to 10-fold when cloned in the sense orientation into the 3'UTR of a CAT expression vector. No block to expression was seen when the fragment was cloned immediately downstream of the poly(A) site, in an intron upstream of the CAT coding sequence, or in an antisense orientation in the 3'UTR. When the same fragment was deleted from a BPV-1 L1 expression vector, a sixfold increase in mRNA levels was seen. Actinomycin D chase experiments using BPV-1 L1 expression vectors indicated that the element does not destabilize cytoplasmic polyadenylated RNA. Therefore, the element must act before the mature mRNA reaches the cytoplasm. The data presented are consistent with effects on nuclear stability and/or inhibition of polyadenylation or nuclear transport.
Furth
P A
PA
Laboratory of Tumor Virus Biology, National Cancer Institute, Bethesda, Maryland 20892.
Baker
C C
CC
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D052688:Bovine papillomavirus 1 / Q000235:genetics*; D002213:Capsid / Q000235:genetics*; D002460:Cell Line; D002872:Chromosome Deletion; D004742:Enhancer Elements, Genetic; D015967:Gene Expression Regulation, Viral; D005814:Genes, Viral; D005822:Genetic Vectors; D051379:Mice; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D016366:Open Reading Frames; D011061:Poly A / Q000235:genetics*; D011401:Promoter Regions, Genetic; D012313:RNA / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics*; D015183:Restriction Mapping; D014162:Transfection
D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger; D011061:Poly A; D012313:RNA