Abstract
How have the branchpoint motifs evolved in organisms of different complexity? Here we identified and examined the consensus motifs (R(1)C(2)T(3)R(4)A(5)Y(6), R: A or G, Y: C or T) of 898 fungal genomes. In Ascomycota unicellular yeasts, the G(4)/A(4) ratio is mostly (98%) below 0.125 but increases sharply in multicellular species by about 40 times on average, and in the more complex Basidiomycota, it increases further by about 7 times. The global G(4) increase is consistent with A(4) to G(4) transitions in evolution. Of the G(4)/A(4)-interacting amino acids of the branchpoint binding protein MSL5 (SF1) and the HSH155 (SF3B1), as well as the 5' splice sites (SS) and U2 snRNA genes, the 5' SS G(3)/A(3) co-vary with the G(4) to some extent. However, corresponding increase of the G(4)-complementary GCAGTA-U2 gene is rare, suggesting wobble-base pairing between the G(4)-containing branchpoint motif and GTAGTA-U2 in most of these species. Interestingly, the G(4)/A(4) ratio correlates well with the abundance of alternative splicing in the two phyla, and G(4) enriched significantly at the alternative 3' SS of genes in RNA metabolism, kinases and membrane proteins. Similar wobble nucleotides also enriched at the 3' SS of multicellular fungi with only thousands of protein-coding genes. Thus, branchpoint motifs have evolved U2-complementarity in unicellular Ascomycota yeasts, but have gradually gained more wobble base-pairing nucleotides in fungi of higher complexity, likely to destabilize branchpoint motif-U2 interaction and/or branchpoint A protrusion for alternative splicing. This implies an important role of relaxing the branchpoint signals in the multicellularity and further complexity of fungi.