Digging into acceptor splice site prediction : an iterative feature selection approach

Feature selection techniques are often used to reduce data dimensionality, increase classification performance, and gain insight into the processes that generated the data. In this paper, we describe an iterative procedure of feature selection and feature construction steps, improving the classification of acceptor splice sites, an important subtask of gene prediction. We show that acceptor prediction can benefit from feature selection, and describe how feature selection techniques can be used to gain new insights in the classification of acceptor sites. This is illustrated by the identification of a new, biologically motivated feature: the AG-scanning feature. The results described in this paper contribute both to the domain of gene prediction, and to research in feature selection techniques, describing a new wrapper based feature weighting method that aids in knowledge discovery when dealing with complex datasets

Prediction of Alternative Splice Sites in Human Genes

This thesis addresses the problem of predicting alternative splice sites in human genes. The most common way to identify alternative splice sites are the use of expressed sequence tags and microarray data. Since genes only produce alternative proteins under certain conditions, these methods are limited to detecting only alternative splice sites in genes whose alternative protein forms are expressed under the tested conditions. I have introduced three multiclass support vector machines that predict upstream and downstream alternative 3’ splice sites, upstream and downstream alternative 5’ splice sites, and the 3’ splice site of skipped and cryptic exons. On a test set extracted from the Alternative Splice Annotation Project database, I was able to correctly classify about 68% of the splice sites in the alternative 3’ set, about 62% of the splice sites in the alternative 5’ set, and about 66% in the exon skipping set

De novo prediction of PTBP1 binding and splicing targets reveals unexpected features of its RNA recognition and function.

The splicing regulator Polypyrimidine Tract Binding Protein (PTBP1) has four RNA binding domains that each binds a short pyrimidine element, allowing recognition of diverse pyrimidine-rich sequences. This variation makes it difficult to evaluate PTBP1 binding to particular sites based on sequence alone and thus to identify target RNAs. Conversely, transcriptome-wide binding assays such as CLIP identify many in vivo targets, but do not provide a quantitative assessment of binding and are informative only for the cells where the analysis is performed. A general method of predicting PTBP1 binding and possible targets in any cell type is needed. We developed computational models that predict the binding and splicing targets of PTBP1. A Hidden Markov Model (HMM), trained on CLIP-seq data, was used to score probable PTBP1 binding sites. Scores from this model are highly correlated (ρ = -0.9) with experimentally determined dissociation constants. Notably, we find that the protein is not strictly pyrimidine specific, as interspersed Guanosine residues are well tolerated within PTBP1 binding sites. This model identifies many previously unrecognized PTBP1 binding sites, and can score PTBP1 binding across the transcriptome in the absence of CLIP data. Using this model to examine the placement of PTBP1 binding sites in controlling splicing, we trained a multinomial logistic model on sets of PTBP1 regulated and unregulated exons. Applying this model to rank exons across the mouse transcriptome identifies known PTBP1 targets and many new exons that were confirmed as PTBP1-repressed by RT-PCR and RNA-seq after PTBP1 depletion. We find that PTBP1 dependent exons are diverse in structure and do not all fit previous descriptions of the placement of PTBP1 binding sites. Our study uncovers new features of RNA recognition and splicing regulation by PTBP1. This approach can be applied to other multi-RRM domain proteins to assess binding site degeneracy and multifactorial splicing regulation

Kernel methods in genomics and computational biology

Support vector machines and kernel methods are increasingly popular in genomics and computational biology, due to their good performance in real-world applications and strong modularity that makes them suitable to a wide range of problems, from the classification of tumors to the automatic annotation of proteins. Their ability to work in high dimension, to process non-vectorial data, and the natural framework they provide to integrate heterogeneous data are particularly relevant to various problems arising in computational biology. In this chapter we survey some of the most prominent applications published so far, highlighting the particular developments in kernel methods triggered by problems in biology, and mention a few promising research directions likely to expand in the future

Fast splice site detection using information content and feature reduction

Background: Accurate identification of splice sites in DNA sequences plays a key role in the prediction of gene structure in eukaryotes. Already many computational methods have been proposed for the detection of splice sites and some of them showed high prediction accuracy. However, most of these methods are limited in terms of their long computation time when applied to whole genome sequence data. Results: In this paper we propose a hybrid algorithm which combines several effective and informative input features with the state of the art support vector machine (SVM). To obtain the input features we employ information content method based on Shannon\u27s information theory, Shapiro\u27s score scheme, and Markovian probabilities. We also use a feature elimination scheme to reduce the less informative features from the input data. Conclusion: In this study we propose a new feature based splice site detection method that shows improved acceptor and donor splice site detection in DNA sequences when the performance is compared with various state of the art and well known method

Using Expressing Sequence Tags to Improve Gene Structure Annotation

Finding all gene structures is a crucial step in obtaining valuable information from genomic sequences. It is still a challenging problem, especially for vertebrate genomes, such as the human genome. Expressed Sequence Tags (ESTs) provide a tremendous resource for determining intron-exon structures. However, they are short and error prone, which prevents existing methods from exploiting EST information efficiently. This dissertation addresses three aspects of using ESTs for gene structure annotation. The first aspect is using ESTs to improve de novo gene prediction. Probability models are introduced for EST alignments to genomic sequence in exons, introns, interknit regions, splice sites and UTRs, representing the EST alignment patterns in these regions. New gene prediction systems were developed by combining the EST alignments with comparative genomics gene prediction systems, such as TWINSCAN and N-SCAN, so that they can predict gene structures more accurately where EST alignments exist without compromising their ability to predict gene structures where no EST exists. The accuracy of TWINSCAN_EST and NSCAN_EST is shown to be substantially better than any existing methods without using full-length cDNA or protein similarity information. The second aspect is using ESTs and de novo gene prediction to guide biology experiments, such as finding full ORF-containing-cDNA clones, which provide the most direct experimental evidence for gene structures. A probability model was introduced to guide experiments by summing over gene structure models consistent with EST alignments. The last aspect is a novel EST-to-genome alignment program called QPAIRAGON to improve the alignment accuracy by using EST sequencing quality values. Gene prediction accuracy can be improved by using this new EST-to-genome alignment program. It can also be used for many other bioinformatics applications, such as SNP finding and alternative splicing site prediction

N-terminal proteomics assisted profiling of the unexplored translation initiation landscape in Arabidopsis thaliana

Proteogenomics is an emerging research field yet lacking a uniform method of analysis. Proteogenomic studies in which N-terminal proteomics and ribosome profiling are combined, suggest that a high number of protein start sites are currently missing in genome annotations. We constructed a proteogenomic pipeline specific for the analysis of N-terminal proteomics data, with the aim of discovering novel translational start sites outside annotated protein coding regions. In summary, unidentified MS/MS spectra were matched to a specific N-terminal peptide library encompassing protein N termini encoded in the Arabidopsis thaliana genome. After a stringent false discovery rate filtering, 117 protein N termini compliant with N-terminal methionine excision specificity and indicative of translation initiation were found. These include N-terminal protein extensions and translation from transposable elements and pseudogenes. Gene prediction provided supporting protein-coding models for approximately half of the protein N termini. Besides the prediction of functional domains (partially) contained within the newly predicted ORFs, further supporting evidence of translation was found in the recently released Araport11 genome re-annotation of Arabidopsis and computational translations of sequences stored in public repositories. Most interestingly, complementary evidence by ribosome profiling was found for 23 protein N termini. Finally, by analyzing protein N-terminal peptides, an in silico analysis demonstrates the applicability of our N-terminal proteogenomics strategy in revealing protein-coding potential in species with well-and poorly-annotated genomes

Discerning Novel Splice Junctions Derived from RNA-Seq Alignment: A Deep Learning Approach

Background: Exon splicing is a regulated cellular process in the transcription of protein-coding genes. Technological advancements and cost reductions in RNA sequencing have made quantitative and qualitative assessments of the transcriptome both possible and widely available. RNA-seq provides unprecedented resolution to identify gene structures and resolve the diversity of splicing variants. However, currently available ab initio aligners are vulnerable to spurious alignments due to random sequence matches and sample-reference genome discordance. As a consequence, a significant set of false positive exon junction predictions would be introduced, which will further confuse downstream analyses of splice variant discovery and abundance estimation. Results: In this work, we present a deep learning based splice junction sequence classifier, named DeepSplice, which employs convolutional neural networks to classify candidate splice junctions. We show (I) DeepSplice outperforms state-of-the-art methods for splice site classification when applied to the popular benchmark dataset HS3D, (II) DeepSplice shows high accuracy for splice junction classification with GENCODE annotation, and (III) the application of DeepSplice to classify putative splice junctions generated by Rail-RNA alignment of 21,504 human RNA-seq data significantly reduces 43 million candidates into around 3 million highly confident novel splice junctions. Conclusions: A model inferred from the sequences of annotated exon junctions that can then classify splice junctions derived from primary RNA-seq data has been implemented. The performance of the model was evaluated and compared through comprehensive benchmarking and testing, indicating a reliable performance and gross usability for classifying novel splice junctions derived from RNA-seq alignment