An element in the bovine papillomavirus late 3' untranslated region reduces polyadenylated cytoplasmic RNA levels

Abstract

Expression of the two bovine papillomavirus type 1 (BPV-1) late genes, Ll 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 Ll 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 6to 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 Ll expression vector, a sixfold increase in mRNA levels was seen. Actinomycin D chase experiments using BPV-1 Ll 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.

The Rev protein of human immunodeficiency virus type 1 counteracts the effect of an AU-rich negative element in the human papillomavirus type 1 late 3' untranslated region

“…Actinomycin D treatment of cells transfected with pCHPV1pA or pCHPV1⌬pA did not reveal a major difference in half-lives of the mRNA transcripts (data not shown). This is in agreement with results of a previous study on the BPV-1 3Ј UTR (21). In that study, Furth and Baker identified inhibitory sequences in the BPV-1 3Ј UTR but did not detect an effect of these sequences on mRNA stability.…”

Insights into the U1 small nuclear ribonucleoprotein complex superfamily

“…Work on human and bovine papillamona virus late gene expression identified an inhibitory element within the 3′ UTR, upstream of the pA site, which contained 5′ ss‐like sequence motifs complementary to the 5′ end of U1 snRNA. Mutations of these motifs abrogated the inhibitory effect, which was restored upon co‐expression of suppressor U1 snRNAs containing complimentary base changes in the 5′ end . These data were further supported by other work investigating pA site selection in the long terminal repeat (LTR) regions that flank the HIV‐1 proviral genome.…”

3′ end mRNA processing: molecular mechanisms and implications for health and disease

“…Polymerase that creates the 3 0 poly(A) tail of mRNAs, also required for the endoribonucleolytic cleavage reaction at some polyadenylation sites, exhibits both nonspecific and CPSF/AAUAAA-dependent polyadenylation activity PAP activity regulated in a cell cycle-dependent manner (Ballantyne et al, 1995;Colgan et al, 1996) U1A inhibits PAP activity on various substrates (i.e., of the U1A mRNA; Gunderson et al, 1994, and of the IgM secretory mRNA; Phillips et al, 2001) U1 snRNP inhibits poly(A) sites and PAP (Furth and Baker, 1991;Gunderson et al, 1998)…”