3'-box-dependent processing of human pre-U1 snRNA requires a combination of RNA and protein co-factors.
Using an in vitro system we have recently shown that the 3' ends of human pre-snRNAs synthesized by RNA polymerase II are produced by RNA processing directed by the snRNA gene-specific 3' box. Towards a complete characterization of this processing reaction we have further investigated the in vitro requirements for proper 3' end formation of pre-U1 snRNA. Here we show that the 5' cap plays a stimulatory role and processing requires creatine phosphate. Our results also indicate that the pre-U1 processing activity is heat sensitive and that an RNA component is required. In addition, the exact sequence adjacent to the 3' box influences the position of the pre-U1 3' end produced in vitro. Interestingly, the processing extract active for 3'-box-dependent processing also contains an activity that converts the 3' end of RNA containing the U1 Sm protein binding site and the 3' terminal stem-loop into the mature form.
Uguen
Patricia
P
Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK. patricia.uguen@ibaic.u-psud.fr
Murphy
Shona
S
D000255:Adenosine Triphosphate / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D003067:Coenzymes / Q000378:metabolism*; D004795:Enzyme Stability; D006367:HeLa Cells; D006358:Hot Temperature; D006801:Humans; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010725:Phosphocreatine / Q000378:metabolism*; D012315:RNA Caps / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional; D012342:RNA, Small Nuclear / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D012261:Ribonucleoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D003067:Coenzymes; D012315:RNA Caps; D012342:RNA, Small Nuclear; D012261:Ribonucleoproteins; C405483:U1 small nuclear RNA; D010725:Phosphocreatine; D000255:Adenosine Triphosphate