RNA Processing in Eukaryotes

 

Summary

Gene expression is extensively regulated at the post-transcriptional level. The fundamental steps of eukaryotic RNA processing have been characterised in great detail, but knowledge of how the disruption of these processes contributes to human disease has only recently begun to emerge. The major aim of this programme is to study the mechanisms for the post-transcriptional regulation of gene expression. Our laboratory studies different aspects of RNA processing, with a particular emphasis on alternative splicing regulation.

 

Key Publications

1. Ellis, J.D.; Lleres, D.; Denegri, M.; Lamond, A.I. and Caceres, J.F. Spatial mapping of splicing factor complexes involved in exon and intron definition. Journal of Cell Biology 181(6):921-934, 2008
   PubMed Abstract
   Comment/ Reviews on this paper
   Robinson, R. A better way to see splicing partners. J Cell Biol 181:875, 2008
2. Michlewski, G.*; Guil, S*.; Semple, C. and Caceres, J.F. Posttranscriptional regulation of miRNAs harboring conserved terminal loops. Molecular Cell 32(3):383-393, 2008 *equal contribution
   PubMed Abstract
3. Michlewski, G.; Sanford, J.R. and Caceres, J.F. The Splicing Factor SF2/ASF regulates translation initiation of cellular mRNAs by enhancing phosphorylation of 4EBP. Molecular Cell 30(3):179-189, 2008 PubMed Abstract
   Comment/ Reviews on this paper
   Bushell, M.; Stoneley, M.; Spriggs, K.A. and Willis, A.E. SF2/ASF TORCs Up Translation. Mol Cell 30:262-263, 2008
   PubMed Abstract
4. Sanford, J.R.; Coutinho, P.; Hackett, J.A.; Wang, X., Ranahan, W. and Caceres, J.F. Identification of nuclear and cytoplasmic mRNA targets for the shuttling protein SF2/ASF. PLoS ONE 3(10):e3369, 2008 PubMed Abstract
5. Guil, S. and Caceres, J.F. The multifunctional RNA-binding protein hnRNP A1 is required for processing of miR-18a. Nat Struct Mol Biol 14:591-596, 2007. PubMed Abstract
   Comment/ Reviews on this paper
   Nielsen A.F., Leuschner, P.J.F. and Martinez, J. Not miR-ly a splicing factor: hnRNP A1 succumbs to microRNA temptation. Nat Struct Mol Bio 14:572-573, 2007
   PubMed Abstract
6. Longman, D.; Plasterk, R.H.; Johnstone,I.L. and Caceres, J.F. Mechanistic insights and identification of two novel factors in the C. elegans NMD pathway. Genes Dev 21:1075-1085, 2007
   PubMed Abstract

 

Collaborations within the Unit

• Professor Nick Hastie
  

 

External Collaborators

• Dr Nicola Gray (MRC HRSU)
• Ben Blencowe (Toronto)
• Iain Johnstone (Glasgow)
  

 

Lab Members

Current lab members involved in this work are:

• Dr Javier F. Caceres - Group leader
  
• Dr Nele Hug - Postdoctoral Scientist
  
• Dr Dasa Longman - Investigator Scientist
  
• Dr Sara Macias Ribela - Postdoctoral Scientist
• Julie Rodor - Career Development Fellow

 

Javier Caceres Lab Link

 

 

We are interested in the trans-acting factors that regulate alternative splicing, such as the SR proteins and hnRNP A/B type of proteins. These proteins have antagonistic activities and their molar ratio influence different modes of alternative splicing in vivo and may represent a mechanism for tissue-specific or developmental regulation of gene expression.

We also study the subcellular distribution of RNA processing factors and how this could be affected by extracellular signals. More recently, we have expanded our interest to RNA processing events that act downstream of pre-mRNA splicing.

 

Purpose

The overall aim of this programme is to increase our understanding of RNA processing in eukaryotes.

 

Approach, Progress and Future Work

We mostly rely on classical molecular biology and cellular biology techniques. Cell culture is widely used and we also are using two animal models, mice and the worm C.elegans.

We have found that hnRNP A1, a factor that has been implicated in alternative splicing and other aspects of RNA processing is required for the processing of an oncogenic microRNA cluster (1, 5). These results underscore a novel role for general RNA binding proteins as auxiliary factors that facilitate the processing of specific miRNAs (Figure 1).

 

Nonsense-mediated mRNA decay (NMD) is a highly conserved surveillance mechanism present in all eukaryotes examined that prevents the synthesis of truncated proteins, by promoting the degradation of mRNAs containing premature translation termination codons (PTCs). It is proposed that NMD modulates the phenotypic outcome of many diseases, since approximately 30% of inherited genetic disorders are caused by frameshift or nonsense mutations that generate premature termination codons. A genome-wide RNAi screen in C.elegans provided evidence for the existence of previously unidentified components of the nonsense-mediated decay (NMD) pathway that are essential for proper embryonic development, which we have termed smgl (for smg lethal). We found that the encoded proteins are conserved throughout evolution and are required for NMD in C. elegans and also in human cells (2).

 

We have created a splicing map of the nucleus. We have used fluorescence resonance energy transfer (FRET) to study wherein the nucleus interactions between splice factors occur (Figure 2). Finally, we demonstrated that a subset of SR proteins shuttle continuously from the nucleus to cytoplasm and are involved in translational regulation; thus, coupling splicing and translation (4).