Two distant upstream regions containing cis-acting signals regulating splicing facilitate 3'-end processing of avian sarcoma virus RNA.
Retroviruses, pararetroviruses, and related retrotransposons generate terminally redundant RNAs by transcription of a template flanked by long terminal repeats in which initiation occurs within the 5' long terminal repeat sequences and 3'-end processing occurs within the 3' long terminal repeat sequences. Processing of avian sarcoma virus RNA is relatively inefficient; approximately 15% of the viral RNA transcripts are read-through products; i.e., they are not processed at the viral poly(A) addition site but at sites in the cellular sequence further downstream. In this report, we show that the efficiency of processing at the viral site is further reduced by deletion of two distant upstream sequences: (i) a 606-nucleotide sequence in the gag gene containing a cis-acting negative regulator of splicing and (ii) a 136-nucleotide sequence spanning the env 3' splice site. The deletion of either or both upstream regions increases the levels of read-through products of both unspliced and spliced viral RNA. In contrast, deletion of the src 3' splice site does not affect the efficiency of processing at the viral poly(A) addition site. The effects on 3'-end processing are not correlated either with distance from the promoter to the poly(A) addition site or with the overall level of viral RNA splicing. Substitution of the avian sarcoma virus poly(A) signal with the simian virus 40 early or late poly(A) signal relieves the requirement for the distant upstream sequences. We propose that cellular factors, which may correspond to splicing factors, bound to the upstream viral sequences may interact with factors bound at the avian sarcoma virus poly(A) signal to stabilize the polyadenylation-cleavage complex and allow for more efficient 3'-end processing.
Miller
J T
JT
Department of Microbiology, University of Iowa, Iowa City 52242.
Stoltzfus
C M
CM
D000818:Animals; D001358:Avian Sarcoma Viruses / Q000235:genetics* / Q000378:metabolism; D015152:Blotting, Northern; D002478:Cells, Cultured; D002642:Chick Embryo; D003609:Dactinomycin / Q000494:pharmacology; D011061:Poly A; D012323:RNA Processing, Post-Transcriptional / Q000235:genetics*; D012367:RNA, Viral / Q000378:metabolism*; D012045:Regulatory Sequences, Nucleic Acid; D014158:Transcription, Genetic; D014162:Transfection
D012367:RNA, Viral; D003609:Dactinomycin; D011061:Poly A