Recognition of the TACTAAC box during mRNA splicing in yeast involves base pairing to the U2-like snRNA

Pseudouridines in spliceosomal snRNAs

“…This recognition involves a base-pairing interaction between the 5' end region of U1 and the 5' splice site of the pre-mRNA. The U2 snRNP then recognizes the branch site through a base-pairing interaction involving the branch site recognition region of U2, thereby bulging out the branch point adenosine of the pre-mRNA (Parker et al, 1987;Zhuang and Weiner, 1989). The joining of U1 and U2 results in the formation of a pre-splicing complex (complex A).…”

Base pairing with U6atac snRNA is required for 5′ splice site activation of U12-dependent introns in vivo

“…Two distinct spliceosomal systems have co-existed in eukaryotic cells since at least the divergence of the plant and animal kingdoms (reviewed in Tarn & Steitz, 1997)+ These two systems act on pairs of mutually incompatible splice sites flanking pre-mRNA introns in eukaryotic nuclear genomes+ The large majority of introns in all known organisms are spliced by a wellstudied pathway requiring the function of the small nuclear RNAs U1, U2, U4, U5, and U6, as well as a large number of additional proteins+ In this pathway, multiple RNA-RNA interactions have been demonstrated to form between the splice site sequences and the snRNAs and between various snRNAs in the spliceosome (reviewed in Nilsen, 1998)+ One of the earliest interactions takes place between the 59 end of U1 snRNA and the 59 splice site via base pairing (Zhuang & Weiner, 1986;Seraphin et al+, 1988;Siliciano & Guthrie, 1988)+ A second base pairing interaction takes place between the sequence in the intron surrounding the site of branching and a region of U2 snRNA (Parker et al+, 1987;Wu & Manley, 1989;)+ Following these initial recognition events, a complex of U4, U5, and U6 snRNPs joins the nascent spliceosome and the combined assemblage undergoes several structural rearrangements+ During this portion of the spliceosome assembly process, the extensive base pairing between U4 snRNA and U6 snRNA is disrupted so that U6 snRNA can participate in base pairing to U2 snRNA (Hausner et al+, 1990;Datta & Weiner, 1991;Wu & Manley, 1991;Madhani & Guthrie, 1992)+ In addition, an adjacent sequence in U6 snRNA forms base pairs to the 59 splice site, which displaces U1 snRNA from the complex (Kandels- Lewis & Seraphin, 1993;Lesser & Guthrie, 1993;Hwang & Cohen, 1996)+ U5 snRNP interacts with exon sequences near the 59 and 39 splice sites, but apparently without substantial sequence specificity (Wyatt et al+, 1992;Sontheimer & Steitz, 1993;Newman, 1997)+ Thus, 59 splice site activation appears to be at least a two-step process in which U1 snRNP, probably in cooperation with additional factors, specifies the 59 splice site followed by U5 and U6 snRNP interactions that activate the site for reaction+ A striking feature of these RNA-RNA interactions is their apparent high degree of conservation throughout eukaryotic phylogeny+ These interactions have been studied in both yeast and human systems in vivo and in vitro by a variety of techniques (see Moore et al+, 1993 andNilsen, 1998 for reviews)+ These studies have demonstrated the stepwise assembly of the spliceosome and the roles of sequence elements in the...…”

ATP-dependent interaction of yeast U5 snRNA loop 1 with the 5′ splice site

“…The spliceosome assembly pathway begins with interaction of the U1 snRNP with the 59 splice site region+ Specifically, the 59 end of the U1 snRNA base pairs at the 59 splice site with protein components of the U1 snRNP stabilizing this interaction (Zhuang & Weiner, 1986;Séraphin et al+, 1988;Siliciano & Guthrie, 1988;Puig et al+, 1999;Zhang & Rosbash, 1999)+ The conserved sequences at the 39 end of the intron, the branchpoint region and 39 splice site, are recognized by a number of protein factors (reviewed in Reed, 2000)+ Following initial recognition of the 59 and 39 ends of the intron, the U2 snRNP binds to the branchpoint region with the U2 snRNA base pairing with the conserved branchpoint sequence (Parker et al+, 1987;Wu & Manley, 1989;Zhuang & Weiner, 1989;Query et al+, 1994)+ Once the U1 and U2 snRNPs interact with the premRNA, a pre-assembled U4/U6+U5 particle interacts with the pre-mRNA and snRNPs already bound to the pre-mRNA+ This results in a number of dynamic and specific RNA and protein rearrangements to form the active spliceosome (Nilsen, 1998;Staley & Guthrie, 1998)+ The RNA rearrangements include U1 base pairing at the 59 splice site being replaced by base pairing of U6 with the conserved intronic sequences and interaction of U5 with exon sequences at the 59 splice site+ In addition, base pairing between U4 and U6 in the U4/U6+U5 particle is dissolved, allowing U6 to form specific base pairing interactions with U2, which re-mains base paired to the branchpoint+ These rearrangements lead to activation of the spliceosome and rapid execution of the two catalytic steps of splicing+ One snRNP that plays a critical role in pre-mRNA splicing is the U5 snRNP+ This is highlighted by U5 being the only snRNP that is a component of both the major (U2) and minor AT-AC (U12) spliceosomes (Tarn & Steitz, 1997)+ Furthermore, the U5 snRNP contains the most evolutionarily conserved splicing factor, Prp8 (Hodges et al+, 1995)+ Prp8 is known to interact with the 59 splice site, 39 splice site, and branchpoint during splicing, as well as with the U5 and U6 snRNAs (Wyatt et al+, 1992;MacMillan et al+, 1994;Teigelkamp et al+, 1995;Umen & Guthrie, 1995;Chiara et al+, 1996Chiara et al+, , 1997Reyes et al+, 1996;Dix et al+, 1998;Vidal et al+, 1999)+ Alleles of Prp8 can also suppress 59 and 39 splice...…”