428 Genome Informatics 14 428–429 (2003) Identifying Potential Regulatory Sequences of Alt

428Genome Informatics14:428–429(2003) Identifying Potential Regulatory Sequences of
Alternative Splicing
Hitomi Itoh1,2Takanori Washio1Masaru Tomita1,2 t00514hi@sfc.keio.ac.jp washy@sfc.keio.ac.jp mt@sfc.keio.ac.jp
1Institute for Advanced Biosciences,Keio University,Tsuruoka,Yamagata997-0035, Japan
2Department of Environmental Information,Keio University,Fujisawa,Kanagawa252-8520,Japan
Keywords:alternative splicing,exonic splicing enhancer(ESE),phylogenetic footprinting
1Introduction
Alternative splicing is an important mechanism that contributes to expanding protein diversity by generating multiple protein isoforms from a single gene.We have previously reported computational approach to infer alternative splicing patterns from Mus musculus full-length cDNA clones and mi-croarray data[4].Although we have predicted a large number of unreported splice variants,general mechanisms that regulate those alternative splicing were yet to be understood.In the present study, we constructed datasets of putative alternatively spliced genes by use of full-length cDNA clones and genomic DNA sequences.Our putative datasets were then evaluated by comparing information con-tents of constitutive splice-sites and those of alternative ing phylogenetic footprinting approach,we identified the potential regulatory sequences of alternative splicing.
2Method and Results
2.1Detection of Putative Alternative Splicing Patterns
We detected putative splice variants using full-length cDNA clones and complete genomic sequences. To construct datasets of putative splice variants and infer complete gene structure,we mapped each full-length cDNA clones to genomic sequences by use of BLAST[1]and SIM4[3].Clones were then grouped into clusters if all the internal regions were correctly mapped.Mapping and clustering results are summarized in Table1.
Table1:Mapping and clustering results.
H.sapiens15,5069,14658,349 6.382,818985
M.musculus49,16219,20087,183 4.548,8791,424
Alternative Splicing Regulatory Sequences429
2.2Identification of Potential Alternative Splicing Regulatory Sequences
As afirst step toward identifying potential regulatory sequences of alternative splicing,we compared the orthologous gene cluster among mammals including mouse,rat,human,cattle,pig,and dog,as shown in Fig.1.We then extracted evolutionary conserved alternative cassette exon and constitutive exon,in order to apply phylogenetic footprinting approach for identifying regulatory sequences.Phy-logenetic footprinting is one of the well-known approaches for the discovery of regulatory sequences in a set of orthologous gene clusters[2].Using this approach,we identified7potential regulatory sequences of alternative splicing.Although experimental verification is lacking,the fact that our ap-proach has identified motifs that have high sequence similarity with known ESE motifs may suggest that these unknown motifs are also functional regulatory element and thus are good novel candidates for functional regulatory sequence.
Figure1:Distribution of mutation rate in evolutionary conserved exons.
3Discussion
Our computational approach identified potential regulatory sequences of alternative splicing.Based on ourfindings,it may be reasonable to suggest that alternative exon tends to have more regulatory sequence for weak splice-sites to be regulated alternatively.The fact that known regulatory sequence motifs were found in our predicted candidate motifs may validate this computational approach and thus indicate that this approach is applicable for identifying potential regulatory sequences of alternative splicing.
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