Mechanism for nuclear pre-mRNA intron splicing

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Pre-mRNA splicing is the process by which introns are removed from pre-mRNA transcripts and the protein coding elements assembled into mature mRNAs before the RNA leaves the nucleus. Pre-mRNA splicing is one of the fundamental processes in constitutive and regulated gene expression in eukaryotes.

  1. Spliceosome assembly pathway

    Unlike Group I and Group II introns, pre-mRNA introns are not able to self-splice without the assistance of trans- acting RNA or protein factors. Splicing of precursor messenger RNA takes place in the spliceosome, a large (~60S) RNA/protein macromolecular machine assembles de novo at each round of splicing.

    The spliceosomal machinery is complex and formed from five ribonucleoprotein subunits, termed uridine-rich small nuclear nucleoproteins (U1, U2, U4, U5, U6 snRNPs), transiently associated to more than 70 non-snRNPs splicing factors (RNA helicases, SR splicing factors, etc...). The detailed mechanism of spliceosome assembly is complex.

  2. Splicing reactions

    The detailed splicing mechanism of pre-mRNA introns is also quite complex. The splicing reaction itself ensues by two consecutive trans-esterification reactions (TER): a bond is in effect transferred from one location to another. Then, the intron is removed and two neighbouring exons are joined together.

    The branch point A residue plays a critical role in the enzymatic reaction.

    1. First trans-esterification reaction

      The first step is a hydrophilic attack. The 2' hydroxyl group of the conserved adenosine within the branching site attacks the conserved guanine of the 5' splicing site at the exon1-intron junction.

      An unusual 2'-5' phosphodiester bond is made between both residues and the exon1-intron junction is cleaved. The products are a 2'-5' phosphodiester RNA lariat structure and a free 3'-OH (leaving group) that arises from the upstream exon.

      A rearrangement of spliceosomal components must follow to permit the second trans-esterification reactions.

    2. Second trans-esterification reaction

      The second step is an other hydrophilic attack. The 3'-OH end of the released exon1 then attacks the scissile phosphodiester bond of the conserved guanine of the 3' splicing site at the intron-exon2 junction. The exon2 is then split.

  3. Ultimate step: Intron release

    This reaction liberates the 3'-OH of the intron resulting in a free lariat and spliced exons. The two exon sequences are joined together, while the intron sequence is released as a lariat structure.

    Intron RNA will be degraded in the nucleus; snRNPs will be recycled.

Last modified: Tue Feb 10 11:19:46 CET 2004