Functional and evolutionary analysis of alternatively spliced genes is consistent with an early eukaryotic origin of alternative splicing

Abstract

Background: Alternative splicing has been reported in various eukaryotic groups including plants, apicomplexans, diatoms, amoebae, animals and fungi. However, whether widespread alternative splicing has evolved independently in the different eukaryotic groups or was inherited from their last common ancestor, and may therefore predate multicellularity, is still unknown. To better understand the origin and evolution of alternative splicing and its usage in diverse organisms, we studied alternative splicing in 12 eukaryotic species, comparing rates of alternative splicing across genes of different functional classes, cellular locations, intron/exon structures and evolutionary origins.

Results: For each species, we find that genes from most functional categories are alternatively spliced. Ancient genes (shared between animals, fungi and plants) show high levels of alternative splicing. Genes with products expressed in the nucleus or plasma membrane are generally more alternatively spliced while those expressed in extracellular location show less alternative splicing. We find a clear correspondence between incidence of alternative splicing and intron number per gene both within and between genomes. In general, we find several similarities in patterns of alternative splicing across these diverse eukaryotes.

Conclusion: Along with previous studies indicating intron-rich genes with weak intron boundary consensus and complex spliceosomes in ancestral organisms, our results suggest that at least a simple form of alternative splicing may already have been present in the unicellular ancestor of plants, fungi and animals. A role for alternative splicing in the evolution of multicellularity then would largely have arisen by co-opting the preexisting process.

Figure 1

Intron/exon numbers and AS frequency. A: Percentage of alternatively spliced genes in different eukaryote genomes vs. the average number of introns per gene. Discontinuous line is an estimated interval for intron density of the ancestor of animals and plants (from 3.5 [18] to 7.0 [15]). B: Frequency of AS versus intron numbers per gene for the 8 species showing relatively high values of AS. Abbreviations: Hsa (Homo sapiens), Mmu (Mus muscullus), Gga (Gallus gallus), Dre (Danio rerio), Cel (Caenorhabditis elegans), Dme (Drosophila melanogaster), Ath (Arabidopsis thaliana), Sce (Saccharomyces cerevisae), Spo (Schizosaccharomyces pombe), Ecu (Encephalitozoon cuniculi), Pfa (Plasmodium falciparum), Cne (Cryptococcus neoformans).

Figure 2

Evolutionary origin of alternatively spliced genes. A: Phylogenetic tree showing the relations between the seven species included in the KOG database and used in this study. PAF (green line) corresponds to the group of KOG's likely appeared before the split of animals, plants and fungi; AF (brown line), KOG's likely appeared in the fungamal ancestor; A (blue line), clusters of orthologous likely appeared in the ancestors of nematodes, insects and vertebrates; LSE's (four wide black lines) correspond to lineage specific expansions of plants, nematodes, insects and vertebrates. B: Percentage of AS for genes according to phylogenetic origin. PAF: ancestral to plants, animals and fungi. AF: ancestor of animals and fungi. A: animals. LSE: lineage specific expansions. Hsa (Homo sapiens), Cel (Caenorhabditis elegans), Dme (Drosophila melanogaster), Ath (Arabidopsis thaliana), Sce (Saccharomyces cerevisae), Spo (Schizosaccharomyces pombe), Ecu (Encephalitozoon cuniculi). Note that in A. thaliana genes can only group into PAF or LSE.

Figure 3

Gene dispensability and alternative splicing. Percentage of alternatively spliced genes according to gene dispensability in evolution. All the gene functions were present in the common ancestor of animals, plants and fungi. Dispensable genes (black): the KOG's to which they belong was lost in at least one of the animal or fungal species included in KOG database. Indispensable genes (grey): KOG's present in the seven studied species.

Figure 4

AS frequency for GO categories for cellular locations. For each category, green/red colored AS frequency indicates that the frequency is higher/lower than the average, with (*) denoting statistical significance. In the "Total" column, the total number of genes of each category is shown (categories represented by less than 35 genes are shown in blue). In parenthesis, for each species, 95% confidence interval for the average of alternatively spliced genes in all Cellular location categories. p-values are given after multiple testing correction. Abbreviations: Hsa (H. sapiens), Mmu (M. musculus), Dme (D. melanogaster), Cel (C. elegans), Cne (C. neoformans), Ath (A. thaliana).

Figure 5

AS frequency for GO categories for molecular function. For each category, green/red colored AS frequency indicates that the frequency is higher/lower than the average, with (*) denoting statistical significance. In the "Total" column, the total number of genes of each category is shown (categories represented by less than 35 genes are shown in blue). In parenthesis, for each species, 95% confidence interval for the average of alternatively spliced genes in all Molecular Function categories. p-values are given after multiple testing correction. Abbreviations: Hsa (H. sapiens), Mmu (M. musculus), Dme (D. melanogaster), Cel (C. elegans), Cne (C. neoformans), Ath (A. thaliana).

Figure 6

AS frequency for GO categories for biological process. For each category, green/red colored AS frequency indicates that the frequency is higher/lower than the average, with (*) denoting statistical significance. In the "Total" column, the total number of genes of each category is shown (categories represented by less than 35 genes are shown in blue). In parenthesis, for each species, 95% confidence interval for the average of alternatively spliced genes in all Biological process categories. p-values are given after multiple testing correction. Abbreviations: Hsa (H. sapiens), Mmu (M. musculus), Dme (D. melanogaster), Cel (C. elegans), Cne (C. neoformans), Ath (A. thaliana).