The effect of intron length on exon creation ratios during the evolution of mammalian genomes

Recent studies report that alternatively spliced exons tend to occur in longer introns, which is attributed to the length constraints for splice site pairing for the two major splicing mechanisms, intron definition versus exon definition. Using genome-wide studies of EST and microarray data from human and mouse, we have analyzed the distribution of various subsets of alternatively spliced exons, based on their inclusion level and evolutionary history, versus increasing intron length. Alternative exons may be included in either a major or minor fraction of all transcripts (known as major-form and minor-form exons, respectively). We find that major-form exons are seven- to eightfold more likely to be contained in short introns (<400 nt) than minor-form exons, which occur preferentially in longer introns. Since minor-form exons are more likely to be novel (approximately 75%), this implied that novel exons arise more frequently in longer introns. To test this hypothesis, we used whole genome alignments to classify exons according to their phylogenetic age. We find that older exons, i.e., exons that are conserved in all mammals, predominate at shorter intron lengths, for both major- and minor-form exons. In contrast, exons that arose recently during primate evolution are more prevalent at longer intron lengths (>1000 nt). This suggests that the observed correlation of longer intron lengths with alternatively spliced exons may be at least partly due to biases in the probability of exon creation, which is higher in long introns.