Figure 1. Consensus sequences of splice sites.

1. Diagram of the 5′ splice site. The 5′ end of the intron, recognized initially by U1 snRNP, is identified by the 9 nt consensus sequence shown. A strong base‐pairing between U1 snRNA and the 5′ splice site increases the likelihood that a splice site is used;
   
2. Diagram of the 3′ splice site.
   

The three elements within approximately 40 nt of the intron's 3′ end are shown here. Because the spliceosomal protein U2AF binds preferentially to pyrimidines, the longer the stretch of uninterrupted pyrimidines in the PPT, the more likely the 3′ splice site will be used.

Figure 2. Splice site definition across the intron or exon.

1. When the intron is small (less than approximately 250 nt) the spliceosome can recognize the splice sites that will be paired across the intron, referring to the intron definition model of splice site recognition;
   
2. When introns are large (greater than approximately 250 nt), splice sites are recognized across the exon.
   

Because splice sites that will be paired have been identified separately (on different exons), an additional step is required to assemble the spliceosome across the intron. This initial definition of splice sites is referred to as “exon definition.”

Figure 3. SREs in action.

1. Enhancement of splicing by ESEs and ISEs occurs through the recruitment of splicing activators that bind to specific RNA sequence elements, thereby recruiting spliceosomal components to nearby splice sites;
   
2. ESSs and ISSs are RNA elements that bind splicing repressors thereby inhibiting or blocking the recruitment of the spliceosomal machinery.
   

The prevalence of SREs throughout spliced transcripts suggests that they play an extensive role in pre‐mRNA splicing.

Figure 4. RNA sequence elements identified in the regulation of SMN pre‐mRNA splicing. The recognition of SMN exon 7 relies on complex interactions between splicing regulatory elements. In this case, a single base change can alter the balance of enhancers and silencers, resulting in different exon 7 inclusion levels. A combinatorial control of exon definition has been proposed to be required for the majority of exons, due to the widespread occurrence of cis‐acting RNA elements within and surrounding exons, including constitutively spliced exons.

Figure 5. The role of RNA secondary structure in splicing. RNA secondary structures can activate splicing by sequestering silencers within stem–loops (left), or silence splicing by concealing a splice site or enhancer sequences (right) within a hairpin. Many alternative splicing events appear to correlate with stable RNA secondary structures [ 73], although the full extent of local and long‐range interactions is unknown.

Figure 6. Summary of cis‐acting RNA elements controlling splice site recognition and exon inclusion. The splice site sequence, intron/exon architecture, RNA secondary structure, enhancer and silencer sequences, as well as links between pre‐mRNA splicing and transcription or polyadenylation, are highlighted. These splicing elements act in concert to mediate the complex regulation of pre‐mRNA splicing.