Analysis of pre-mRNA splicing in yeast

Abstract

Abstract Many simple but powerful techniques have been developed to investigate gene expression in Saccharomyces cerevisiae, and a number of these have been employed in the analysis of RNA splicing. Test substrates can be transcribed from expression cassettes introduced into yeast by transformation or transplacement, and splicing can be monitored by RNA analysis or assay of a suitable reporter gene product. Gene disruption and inducible expression systems have been invaluable for investigating the roles of components of the splicing machinery in yeast. Many of the genes for yeast splicing factors have recently been isolated by molecular cloning, which is facilitated by the compact nature of the Saccharomyces genome and the fact that these genes are present as single copies. Splicing of yeast mRNA precursors can also be studied in vitro, since for Saccharomyces a simple method has been developed for making cell-free extracts capable of splicing synthetic pre-mRNA substrates. Such studies have shown that the splicing pathway in yeast is very similar to that of higher eukaryotes and that there is considerable conservation of structure and function between splicing factors in yeast and mammalian cells. However, yeast is an extremely valuable system for the detailed analysis of splicing since it allows some approaches which are not possible with mammalian cells, particularly the genetic selection and isolation of splicing mutants.