Efficient polyadenylation of Rous sarcoma virus RNA requires the negative regulator of splicing element.
Rous sarcoma virus pre-mRNA contains an element known as the negative regulator of splicing (NRS) that acts to inhibit viral RNA splicing. The NRS binds serine/arginine-rich (SR) proteins, hnRNP H and the U1/U11 snRNPs, and appears to inhibit splicing by acting as a decoy 5' splice site. Deletions within the gag gene that encompass the NRS also lead to increased read-through past the viral polyadenylation site, suggesting a role for the NRS in promoting polyadenylation. Using NRS-specific deletions and mutations, we show here that a polyadenylation stimulatory activity maps directly to the NRS and is most likely dependent upon SR proteins and U1 and/or U11 snRNP. hnRNP H does not appear to mediate splicing control or stimulate RSV polyadenylation, since viral RNAs containing hnRNP H-specific mutations were spliced and polyadenylated normally. However, the ability of hnRNP H mutations to suppress the read-through caused by an SR protein mutation suggests the potential for hnRNP H to antagonize polyadenylation. Interestingly, disruption of splicing control closely correlated with increased read-through, indicating that a functional NRS is necessary for efficient RSV polyadenylation rather than binding of an individual factor. We propose a model in which the NRS serves to enhance polyadenylation of RSV unspliced RNA in a process analogous to the stimulation of cellular pre-mRNA polyadenylation by splicing complexes.
Fogel
Brent L
BL
Medical College of Wisconsin, Department of Microbiology and Molecular Genetics, 8701 Watertown Plank Road, Milwaukee, WI 53226, USA.
McNally
Lisa M
LM
McNally
Mark T
MT
D000818:Animals; D001358:Avian Sarcoma Viruses / Q000235:genetics*; D001665:Binding Sites; D002642:Chick Embryo; D005347:Fibroblasts; D005814:Genes, Viral / Q000235:genetics; D006367:HeLa Cells; D034541:Heterogeneous-Nuclear Ribonucleoprotein Group F-H; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D006801:Humans; D008957:Models, Genetic; D009154:Mutation / Q000235:genetics; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D023282:Nuclease Protection Assays; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D026723:Polyadenylation / Q000235:genetics*; D011485:Protein Binding; D012322:RNA Precursors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012326:RNA Splicing / Q000235:genetics*; D012367:RNA, Viral / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics*; D017412:Ribonucleoprotein, U1 Small Nuclear / Q000378:metabolism; D012261:Ribonucleoproteins / Q000235:genetics / Q000378:metabolism; D000068103:Serine-Arginine Splicing Factors; D020798:Two-Hybrid System Techniques
D034541:Heterogeneous-Nuclear Ribonucleoprotein Group F-H; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D009687:Nuclear Proteins; D010750:Phosphoproteins; D012322:RNA Precursors; D012367:RNA, Viral; D016601:RNA-Binding Proteins; D017412:Ribonucleoprotein, U1 Small Nuclear; D012261:Ribonucleoproteins; D000068103:Serine-Arginine Splicing Factors