Conservation of mRNA quality control factor Ski7 and its diversification through changes in alternative splicing and gene duplication.

Eukaryotes maintain fidelity of gene expression by preferential degradation of aberrant mRNAs that arise by errors in RNA processing reactions. In Saccharomyces cerevisiae, Ski7 plays an important role in this mRNA quality control by mediating mRNA degradation by the RNA exosome. Ski7 was initially thought to be restricted to Saccharomyces cerevisiae and close relatives because the SKI7 gene and its paralog HBS1 arose by whole genome duplication (WGD) in a recent ancestor. We have recently shown that the preduplication gene was alternatively spliced and that Ski7 function predates WGD. Here, we use transcriptome analysis of diverse eukaryotes to show that diverse eukaryotes use alternative splicing of SKI7/HBS1 to encode two proteins. Although alternative splicing affects the same intrinsically disordered region of the protein, the pattern of splice site usage varies. This alternative splicing event arose in an early eukaryote that is a common ancestor of plants, animals, and fungi. Remarkably, through changes in alternative splicing and gene duplication, the Ski7 protein has diversified such that different species express one of four distinct Ski7-like proteins. We also show experimentally that the Saccharomyces cerevisiae SKI7 gene has undergone multiple changes that are incompatible with the Hbs1 function and may also have undergone additional changes to optimize mRNA quality control. The combination of transcriptome analysis in diverse eukaryotes and genetic analysis in yeast clarifies the mechanism by which a Ski7-like protein is expressed across eukaryotes and provides a unique view of changes in alternative splicing patterns of one gene over long evolutionary time.

Alternative splicing and subfunctionalization generates functional diversity in fungal proteomes.

Alternative splicing is commonly used by the Metazoa to generate more than one protein from a gene. However, such diversification of the proteome by alternative splicing is much rarer in fungi. We describe here an ancient fungal alternative splicing event in which these two proteins are generated from a single alternatively spliced ancestral SKI7/HBS1 gene retained in many species in both the Ascomycota and Basidiomycota. While the ability to express two proteins from a single SKI7/HBS1 gene is conserved in many fungi, the exact mechanism by which they achieve this varies. The alternative splicing was lost in Saccharomyces cerevisiae following the whole-genome duplication event as these two genes subfunctionalized into the present functionally distinct HBS1 and SKI7 genes. When expressed in yeast, the single gene from Lachancea kluyveri generates two functionally distinct proteins. Expression of one of these proteins complements hbs1, but not ski7 mutations, while the other protein complements ski7, but not hbs1. This is the first known case of subfunctionalization by loss of alternative splicing in yeast. By coincidence, the ancestral alternatively spliced gene was also duplicated in Schizosaccharomyces pombe with subsequent subfunctionalization and loss of splicing. Similar subfunctionalization by loss of alternative splicing in fungi also explains the presence of two PTC7 genes in the budding yeast Tetrapisispora blattae, suggesting that this is a common mechanism to preserve duplicate alternatively spliced genes.