The spliceosome is a hugely massive complex that catalyzes excision of introns and ligation of exons.
In addition to many non-snRNP subunits, the spliceosome consists of 5 small nuclear ribonucleoproteins (snRNPs) that assemble stepwise onto each intron. The snRNPs are small and are composed of: a small nuclear RNA (snRNA) with a specific sequence and secondary structure; and some common and some specific proteins.
The five snRNPs of the spliceosome: U1, U2, U4 and U5 are transcribed by RNAP II and have a TriMethyl G Cap; U6 is transcribed by RNAP III and has an unusual Cap. In the U4/U6 Di-snRNP and the U4/U5/U6 Tri-snRNP, the U4 and U6 snRNAs are base-paired to each other. This interaction is later disrupted in the formation of the active spliceosome.
Remember that the splice sites do not always perfectly match the consensus sequences. Thus, the base-pairing interactions between the snRNAs and the pre-mRNA are not always the same. The interactions of U1 with the 5’ splice site and U2 with the branchpoint were proven by creating mutant splice sites that disrupted pairing with the snRNA and inhibited splicing. Compensating mutations in the snRNA that restored complementarity (base-pairing) with the splice site restored splicing.
|U1||U1 is the first snRNP to bind the pre-mRNA; uses its ssRNA to base-pair to the 5’ splice site.|
|U2AF||U2 Auxilliary Factor (U2AF) binds the 3' splice site's polypyrimidine tract and Ag, then helps U2 bind to the branchpoint.|
|U2||Completing the ATP-dependent pre-spliceosomal A Complex, U2 binds to the branchpoint via RNA/RNA base-pairs to create a bulged A residue.|
|U4, U5 & U6||The pre-spliceosome becomes a full spliceosome via addition of the U4/U5/U6 Tri-snRNP. This step also requires ATP hydrolysis. U6 base-pairs to U2 snRNA now, instead of to the U4 snRNP.|
|Detachment||In an ATP-dependent process, U1 and U4 detach and U2, U5 and U6 are rearranged.|
|All Done!||In the mature catalytically active spliceosome, the U1 and U4 snRNPs are absent, and the U2 and U6 snRNPs are basepaired to each other. The U2, U5 and U6 snRNAs, and the large spliceosomal protein Prp8 all make very specific contacts with the splice sites.|