Most metazoan genes have multiple exons that must be carefully excised (from the pre-mRNA) and then ligated (to form the mRNA). This RNA splicing occurs in the nucleus, and upon its completion the mRNA is exported to the cytoplasm for translation. Exon definition complexes and spliceosomes begin to assemble during transcription. Some of these complexes interact with RNAP II's CTD, and some introns are thus excised before transcription has even terminated.
There is no apparent order in which introns get spliced). Introns do no excise in any particular order, and active transcription (or its termination) is not needed for splicing to occur. However, the rate of transcription elongation through an intron can strongly affect what splice sites are chosen and thus indirectly couple transcription and splicing. Exon definition complexes and spliceosomes begin assembling during RNA synthesis; interaction with RNAP II's CTD leads to excision of some introns before transcription has even terminated).
Whether a gene has one or many introns, each is excised via the same two isoenergetic transesterification reactions:
- First transesterification Within the intron, the 2' hydroxyl of the branchpoint A attacks the 5'-end phosphate, releasing the 5' exon and forming an intron lariat. The new phosphodiester bond of the A nucleotide makes it a branched nucleotide bound to its attack-ee and two adjacent bases. The branched nucleotide is a most peculiar phenomenon.
- Second transesterification The 3' hydroxyl of the detached 5' exon attacks the phosphate at the 3' end of the intron. This causes release of the intron lariat and ligation of the exons.