Poster abstracts
Poster number 12 submitted by Caleb Embree
Disruption of pre-mRNA Splicing in Spliceosomapathies is Likely to Impact Downstream mRNA Quality Control Pathways
Caleb Embree (Department of Molecular Genetics, Center for RNA Biology), Andreas Stephanou (Department of Molecular Genetics), Guramrit Singh (Department of Molecular Genetics, Center for RNA Biology)
Abstract:
To create protein molecules within a eukaryotic cell, information encoded in DNA is first transcribed into precursor messenger RNAs (pre-mRNA), from where extraneous sequences are removed via pre-mRNA splicing to make the final mRNA that will act as template for protein synthesis. pre-mRNA splicing is carried out by a large and dynamic ribonucleoprotein machine known as the spliceosome (1). The spliceosome also decorates mRNA exon junctions with the exon junction complex (EJC), which is critical for proper mRNA expression including monitoring of premature translation termination by the nonsense mediated mRNA decay (NMD) pathway (2). Mutations in components of the spliceosome cause a number of disorders, most commonly affecting cranio-facial and neurological development in humans (3). To determine the effect of spliceosomal protein depletion on NMD, we analyzed publicly available spliceosome knockdown (KD) RNA-seq datasets. When components of the spliceosome involved in the catalytic steps of splicing are depleted, naturally occurring NMD targets are upregulated, indicating a defective NMD pathway. Such a defect can either be caused by an increased burden on the NMD machinery due to overproduction of NMD targeted mRNAs due to widespread mis-splicing or result from compromised EJC deposition. To test these two possibilities, we have focused on EFTUD2, a GTPase that is critical for catalytic activation of the spliceosome (4). Notably, mutations in human EFTUD2 leads to a rare cranio-facial disorder known as Mandibulofacial Dysostosis, Guion-Almedia type (MFDGA) (5). All the pathogenic mutations tested compromise interaction with BRR2, EFTUD2’s primary interactor within the spliceosome, whereas EFTUD2 truncating mutations also eliminate its interaction with the EJC and reduce protein stability. Together these preliminary data suggest that the impact on NMD upon spliceosome disruption may result from a combination of over-production of NMD targets and impaired EJC deposition at the same time.
Overall, where our work provides the first look at the molecular changes that underpin MFDGA, it also provides new insights into the multi-layered relationship between pre-mRNA splicing and RNA quality control that is likely to be impacted in a variety of spliceosomopathies.
References:
1. Wilkinson, M. E., Charenton, C., and Nagai, K. (2020) RNA Splicing by the Spliceosome. Annu. Rev. Biochem. 89, 1-1.30
2. Embree, C. M., Abu-Alhasan, R., and Singh, G. (2022) Features and factors that dictate if terminating ribosomes cause or counteract nonsense-mediated mRNA decay. J. Biol. Chem. 10.1016/j.jbc.2022.102592
3. Griffin, C., and Saint-Jeannet, J.-P. (2020) Spliceosomopathies: Diseases and mechanisms. Dev. Dyn. 249, 1038–1046
4. Bartels, C., et al. (2002) The ribosomal translocase homologue Snu114p is involved in unwinding U4/U6 RNA during activation of the spliceosome. EMBO Rep. 3, 875–880
5. Guion-Almeida, et al. (2006) A new syndrome with growth and mental retardation, mandibulofacial dysostosis, microcephaly, and cleft palate. Clin. Dysmorphol. 15, 171–174
Keywords: Splicing, NMD, RNA regulation