title: Direct interaction of the U1 snRNP-A protein with the upstream efficiency element of the SV40 late polyadenylation signal.
abstract: An integral component of the splicing machinery, the U1 snRNP, is here implicated in the efficient polyadenylation of SV40 late mRNAs. This occurs as a result of an interaction between U1 snRNP-A protein and the upstream efficiency element (USE) of the polyadenylation signal. UV cross-linking and immunoprecipitation demonstrate that this interaction can occur while U1 snRNP-A protein is simultaneously bound to U1 RNA as part of the snRNP. The target RNA of the first RRM (RRM1) has been shown previously to be the second stem-loop of U1 RNA. We have found that a target for the second RRM (RRM2) is within the AUUUGURA motifs of the USE of the SV40 late polyadenylation signal. RNA substrates containing the wild-type USE efficiently bind to U1 snRNP-A protein, whereas substrates fail to bind when motifs of the USE were replaced by linker sequences. The addition of an oligoribonucleotide containing a USE motif to an in vitro polyadenylation reaction inhibits polyadenylation of a substrate representing the SV40 late polyadenylation signal, whereas a mutant oligoribonucleotide, a nonspecific oligoribonucleotide, and an oligoribonucleotide containing the U1 RNA-binding site had much reduced or no inhibitory effects. In addition, antibodies to bacterially produced, purified U1 snRNP-A protein specifically inhibit in vitro polyadenylation of the SV40 late substrate. These data suggest that the U1 snRNP-A protein performs an important role in polyadenylation through interaction with the USE. Because this interaction can occur when U1 snRNP-A protein is part of the U1 snRNP, our data provide evidence to support a link between the processes of splicing and polyadenylation, as suggested by the exon definition model.
authors.affiliation: Department of Microbiology, School of Medicine, University of Pennsylvania, Philadelphia 19104-6142.
authors.forename: C S
authors.initials: CS
authors.lastname: Lutz
authors.forename: J C
authors.initials: JC
authors.lastname: Alwine
mesh_terms: D001483:Base Sequence; D008969:Molecular Sequence Data; D011061:Poly A / Q000378:metabolism; D011233:Precipitin Tests; D012333:RNA, Messenger / Q000378:metabolism; D017412:Ribonucleoprotein, U1 Small Nuclear / Q000378:metabolism*; D013539:Simian virus 40 / Q000235:genetics*; D014158:Transcription, Genetic; D014466:Ultraviolet Rays
chemical_list: D012333:RNA, Messenger; D017412:Ribonucleoprotein, U1 Small Nuclear; D011061:Poly A