Alternative splicing creates transcriptome diversification, possibly leading to speciation. A large fraction of the protein-coding genes of multicellular organisms are alternatively spliced, although no regulated splicing has been detected in unicellular eukaryotes such as yeasts. A comparative analysis of unicellular and multicellular eukaryotic 5' splice sites has revealed important differences - the plasticity of the 5' splice sites of multicellular eukaryotes means that these sites can be used in both constitutive and alternative splicing, and for the regulation of the inclusion/skipping ratio in alternative splicing. So, alternative splicing might have originated as a result of relaxation of the 5' splice site recognition in organisms that originally could support only constitutive splicing.