Post Doctoral Researcher

Andrea Pawellek

a.pawellek@dundee.ac.uk
Andrea Pawellek
Germany
Andrea Pawellek

Project Description


Pre-mRNA splicing is an important step in gene expression. However in contrast to other steps involved in gene expression, including transcription and translation, very few well characterized chemical inhibitors are available with which to dissect or study the splicing process particularly in vivo. Therefore, the identification of specific and selective splicing inhibitors/modifiers would not only be extremely valuable for research purposes as biotools, but also potentially useful for therapeutic applications.

To date only a few natural compounds and their synthetic derivatives have been identified as general splicing inhibitors. In addition, several other natural compounds, derived either from extracts of plants, or microbes, have been reported either to inhibit splicing in vitro, or to change splicing of certain transcripts in cells. Our aim is to identify new small molecules that modulate splicing in vitro and in cells. We conducted a high throughput screen of 71,504 small, drug-like chemical compounds by using a recently published high throughput in vitro splicing assay (1). Potential hits were verified by radioactive and RT-PCR based in vitro splicing assays and replicated between nuclear extracts from different human cell types. All compounds that tested positive for splicing inhibition in vitro were further tested in HeLa cells for their ability to inhibit splicing of endogenous genes in vivo. 10 new compounds were identified that modify splicing in vitro and inhibit pre-mRNA in cells. A further screen of ca. 100 derivatives of one of the new splicing inhibitors identified an additional 15 compounds that act as general splicing modulators.



Figure
Figure
Schematic representation of the non-radioactive in vitro splicing assay used for low throughput screening (A) and of the characterization of splicing modulators in cells (B).
References

1. Samatov T et al., ChemBioChem Volume 13, Issue 5, pages 640–644, March 19, 2012

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