The U1 snRNP protein U1C recognizes the 5' splice site in the absence of base pairing.
Splicing of precursor messenger RNA takes place in the spliceosome, a large RNA/protein macromolecular machine. Spliceosome assembly occurs in an ordered pathway in vitro and is conserved between yeast and mammalian systems. The earliest step is commitment complex formation in yeast or E complex formation in mammals, which engages the pre-mRNA in the splicing pathway and involves interactions between U1 small nuclear ribonucleoprotein (snRNP) and the pre-mRNA 5' splice site. Complex formation depends on highly conserved base pairing between the 5' splice site and the 5' end of U1 snRNA, both in vivo and in vitro. U1 snRNP proteins also contribute to U1 snRNP activity. Here we show that U1 snRNP lacking the 5' end of its snRNA retains 5'-splice-site sequence specificity. We also show that recombinant yeast U1C protein, a U1 snRNP protein, selects a 5'-splice-site-like sequence in which the first four nucleotides, GUAU, are identical to the first four nucleotides of the yeast 5'-splice-site consensus sequence. We propose that a U1C 5'-splice-site interaction precedes pre-mRNA/U1 snRNA base pairing and is the earliest step in the splicing pathway.
Du
Hansen
H
Howard Hughes Medical Institute, Department of Biology, MS008, Brandeis University, 415 South Street, Waltham, Massachusetts 02454, USA.
Rosbash
Michael
M
D000818:Animals; D020029:Base Pairing; D001483:Base Sequence; D002457:Cell Extracts; D005363:Ficusin / Q000378:metabolism; D009690:Nucleic Acid Conformation; D012322:RNA Precursors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D022821:RNA Splice Sites / Q000235:genetics*; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012342:RNA, Small Nuclear / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017412:Ribonucleoprotein, U1 Small Nuclear / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017411:Ribonucleoproteins, Small Nuclear / Q000378:metabolism*; D013379:Substrate Specificity; D013696:Temperature; D015003:Yeasts / Q000166:cytology / Q000235:genetics
D002457:Cell Extracts; D012322:RNA Precursors; D022821:RNA Splice Sites; D012333:RNA, Messenger; D012342:RNA, Small Nuclear; D017412:Ribonucleoprotein, U1 Small Nuclear; D017411:Ribonucleoproteins, Small Nuclear; C405483:U1 small nuclear RNA; D005363:Ficusin