7.5 Alternative Splicing in ammals 137the cofactor attacks the 5′ end of the intron, resulting in the first transesterifica-
tion reaction. The free 3′ hydroxyl of the first exon thus generated then attacks
the junction between intron and second exon, leading to the second transesteri-
fication step. Consequently, the intron is released as a linear molecule that
circularizes later [41].
Group II introns share the same catalytic mechanism as nuclear pre-mRNA
introns excised by the spliceosome with the first nucleophilic attack of a branch
point adenosine, resulting in lariat formation of the intron (see Section 7.2.2)
[42]. Interestingly, recent data indicate that the U6 snRNA of the spliceosome
catalyzes both splicing steps by positioning divalent metal ions so that they sta-
bilize the leaving group during each reaction. Notably, all ligands of the catalyti-
cally active metal ions in the U6 snRNA correspond to ligands observed to
position catalytically active divalent metals in the crystal structures of group II
intron RNAs [43]. This agreement indicates that group II introns and the spli-
ceosome share common catalytic mechanisms and probably common evolution-
ary origins [44]. It also suggests that splicing evolved from an autocatalytic
reaction inherent to an individual RNA molecule [45]. As splicing became more
complex, proteins started to play a more important role. Importantly, the simi-
larities between the catalytic core of the group II intron and the U6 snRNA sup-
port the hypothesis that spliceosomal introns in eukaryotes developed out of
group II self-splicing introns [46].
7.4.2 tRNA Splicing
The splicing of tRNAs in archaea and eukarya is the only example of intron
removal that does not involve transesterification, but instead successive cleavage
and ligation reactions. tRNAs contain a single intron located one nucleotide next
to the anticodon. These introns are short (14–60 nucleotides) and have no con-
sensus sequence. They are recognized by an endonuclease that detects a com-
mon secondary structure of the tRNA rather than a sequence element. It cleaves
both ends of the intron generating two tRNA halves that are subsequently joined
by an RNA ligase [47].
7.5 Alternative Splicing in Mammals
7.5.1 Different Mechanisms of Alternative Splicing
Alternative splicing affects 95% of all human genes [48, 49] and produces multi-
ple mRNA molecules from a single gene. The resulting proteomic diversity
is important for many different cellular processes, including cell growth and
differentiation [13].
Alternative splicing events can be divided into four major categories: inclusion
and exclusion of (cassette) exons, the usage of alternative 5′ or 3′ splice sites, and
the retention of entire introns (see Figure 7.2). Of these, the cassette exon type
accounts for approximately one third of all alternative splicing events in humans
[50]. Cassette exons are either fully included or excluded in the mature mRNA.