Implicit alternative splicing prediction
An alternative approach is to predict multiple overlapping gene structures, or a single gene structure overlapping existing alternative annotation. Explicit features of alternative splicing are not scored, but by virtue of having multiple overlapping high scoring gene structures, alternative splicing is implied. One method sampled paths [19] in the generalized hidden Markov Model (GHMM) of the single isoform gene finder SLAM [20]. Re-occurring overlapping high scoring parses were reported as candidates for alternative splicing. Another approach is to find an exon splicing pattern with the highest scoring alignment to profile hidden Markov models (profile-HMMs) [21]. The human genome was searched for cassette exons and intron retention events using a reference annotation [22]. Predicted gene structures with scores exceeding the reference gene structure were inferred to be examples of alternative splicing.
The work most similar to the model introduced in this article is the pair-HMM UNCOVER [23], which finds exons in sequence annotated as introns and was tested on human/mouse intron pairs. Unlike the cassette exon classification methods [15-17], models were trained using examples of protein coding exons without explicitly distinguishing between constitutive exons and cassette exons. Since the input sequence is assumed to be an intron, predicted exons are inferred to be alternatively spliced.
The method presented in this article extends the GHMMs used in single isoform gene finding [24] to explicitly model features of alternative and constitutive exons. The features of the explicit alternative splicing prediction methods: k-mer counts, exon lengths, and sequence conservation are used to predict multiple splice sites and intron retention events along with cassette exons and constitutive exons. Cross-species sequence conservation is incorporated using components of the single isoform phylogenetic HMM gene finders [25-27]. The phylogenetic shadowing principle is used to assume a multiple sequence alignment can be obtained from closely related species [28]. In contrast, the pair-HMM method simultaneously predicts a pairwise alignment and the exon structure making it potentially better suited to incorporate a difficult to align, more distantly related organism. Conservation from greater evolutionary distances may improve discriminative power in identify functional nucleotides, but with the potential trade off of detecting a smaller set of conserved alternative splicing events [29].
The remainder of the article describes our computational prediction model and reports on prediction accuracy in Drosophila melanogaster.