Characterization of nonsense mediated mRNA decay in Schizosaccharomyces pombe

Wen, Jikai (2010). Characterization of nonsense mediated mRNA decay in Schizosaccharomyces pombe. University of Birmingham. Ph.D.

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Abstract

Nonsense mediated mRNA decay (NMD) is a translation-coupled process that preferentially destroys mRNAs harboring premature translation termination codons (PTCs). In mammalian cells, NMD is linked to pre-mRNA splicing. Typically, PTCs elicit strong NMD only if positioned upstream of at least one intron. The exon junction complex (EJC) is believed to mediate the link between splicing and NMD. However, recent studies have questioned the importance of splicing and the EJC in NMD and instead they have proposed that, from yeast to mammalian cells, NMD is mostly determined by the distance of the PTC from the 3’ end. In this study, to investigate the link between pre-mRNA splicing and NMD, I used the fission yeast Schizosaccharomyces pombe. Many genes carry introns in fission yeast and unlike in Saccharomyces cerevisiae, the genome encodes for proteins homologous to EJC components. S. pombe is a powerful model organism which is easily genetically manipulated and we envisaged that studying NMD in this organism should facilitate the understanding of the molecular mechanism and in particular the link between NMD and splicing. During my PhD research, I have developed a versatile gene reporter system to study NMD; with it I discovered that splicing strongly enhances NMD in fission yeast and, surprisingly, that the EJC does not appear to be required. Unexpectedly, I found that splicing enhances NMD when the intron is either before or after the PTC, and furthermore, it does so only when the intron is close to the PTC. These observations suggest that the effect of splicing on NMD is direct and not a secondary consequence of splicing enhancing translation. Splicing is not an absolute requirement for NMD; I found that PTCs located early in the coding region could induce NMD even in the absence of a nearby intron. However, against the prediction of current models, I found no strong correlation with the distance of the PTC from the 3’ end. In summary, during my PhD a versatile system has been developed to study NMD in fission yeast and what I have observed challenges current NMD models and provides new mechanistic insights into NMD.

Type of Work:

Thesis (Doctorates > Ph.D.)

Award Type:

Doctorates > Ph.D.

Supervisor(s):