Splicing of a divergent subclass of AT-AC introns requires the major spliceosomal snRNAs.
AT-AC introns constitute a minor class of eukaryotic pre-mRNA introns, characterized by 5'-AT and AC-3' boundaries, in contrast to the 5'-GT and AG-3' boundaries of the much more prevalent conventional introns. In addition to the AT-AC borders, most known AT-AC introns have highly conserved 5' splice site and branch site sequence elements of 7-8 nt. Intron 6 of the nucleolar P120 gene and intron 2 of the SCN4A voltage-gated skeletal muscle sodium channel are AT-AC introns that have been shown recently to be processed via a unique splicing pathway involving several minor U snRNAs. Interestingly, intron 21 of the same SCN4A gene and the corresponding intron 25 of the SCN5A cardiac muscle sodium channel gene also have 5'-AT and AC-3' boundaries, but they have divergent 5' splice site and presumptive branch site sequences. Here, we report the accurate in vitro processing of these two divergent AT-AC introns and show that they belong to a functionally distinct subclass of AT-AC introns. Splicing of these introns does not require U12, U4atac, and U6atac snRNAs, but instead requires the major spliceosomal snRNAs U1, U2, U4, U5, and U6. Previous studies showed that G --> A mutation at the first position and G --> C mutation at the last position of a conventional yeast or mammalian GT-AG intron suppress each other in vivo, suggesting that the first and last bases participate in an essential non-Watson-Crick interaction. Our results show that such introns, hereafter termed AT-AC II introns, occur naturally and are spliced by a mechanism distinct from that responsible for processing of the apparently more common AT-AC I introns.
Wu
Q
Q
Cold Spring Harbor Laboratory, New York 11724, USA.
Krainer
A R
AR
D001483:Base Sequence; D005075:Biological Evolution; D002467:Cell Nucleus / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007438:Introns; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000235:genetics; D062553:NAV1.4 Voltage-Gated Sodium Channel; D062554:NAV1.5 Voltage-Gated Sodium Channel; D009687:Nuclear Proteins / Q000235:genetics; D012322:RNA Precursors / Q000378:metabolism*; D012326:RNA Splicing; D012333:RNA, Messenger / Q000378:metabolism*; D012342:RNA, Small Nuclear / Q000378:metabolism*; D017413:Ribonucleoprotein, U2 Small Nuclear; D012689:Sequence Homology, Nucleic Acid; D015222:Sodium Channels / Q000235:genetics; D017381:Spliceosomes / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism; D043325:Xylan Endo-1,3-beta-Xylosidase; D014995:Xylosidases / Q000235:genetics; D012359:tRNA Methyltransferases
D009124:Muscle Proteins; D062553:NAV1.4 Voltage-Gated Sodium Channel; D062554:NAV1.5 Voltage-Gated Sodium Channel; D009687:Nuclear Proteins; D012322:RNA Precursors; D012333:RNA, Messenger; D012342:RNA, Small Nuclear; D017413:Ribonucleoprotein, U2 Small Nuclear; C106251:SCN4A protein, human; C568320:SCN5A protein, human; D015222:Sodium Channels; C096986:U12 small nuclear RNA; C065028:NOP2 protein, human; D012359:tRNA Methyltransferases; D014995:Xylosidases; D043325:Xylan Endo-1,3-beta-Xylosidase