The U5 and U6 Small Nuclear RNAs as Active Site Components of the Spliceosome
0303 health sciences
Binding Sites
Base Sequence
RNA Splicing
Molecular Sequence Data
Thiouridine
Exons
Catalysis
Adenoviridae
03 medical and health sciences
RNA, Small Nuclear
RNA Precursors
Spliceosomes
Nucleic Acid Conformation
RNA, Viral
DOI:
10.1126/science.8266094
Publication Date:
2006-10-05T23:05:09Z
AUTHORS (2)
ABSTRACT
Five small nuclear RNAs (U1, U2, U4, U5, and U6) participate in precursor messenger RNA (pre-mRNA) splicing. To probe their interactions within the active center of the mammalian spliceosome, substrates containing a single photoactivatable 4-thiouridine residue adjacent to either splice site were synthesized, and crosslinks were induced during the course of in vitro splicing. An invariant loop sequence in U5 small nuclear RNA contacts exon 1 before and after the first step of splicing because a crosslink between U5 and the last residue of exon 1 appeared in the pre-mRNA and then in the cutoff exon 1 intermediate. Both of these crosslinked species could undergo subsequent splicing, indicating that the crosslinks reflect a functional interaction that is maintained through both reaction steps. The same U5 loop aligns the two exons for ligation since the first residue of exon 2 also became crosslinked to U5 in the lariat intermediate. An invariant sequence in U6 RNA became crosslinked to the conserved second position of the intron within both the lariat intermediate and the lariat intron product. On the basis of these results, several conformational arrangements of small nuclear RNAs within the spliceosomal active center can be distinguished, and additional mechanistic parallels between the spliceosome and self-splicing introns can be drawn.
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