Journal article

A systems view of spliceosomal assembly and branchpoints with iCLIP

Michael Briese, Nejc Haberman, Christopher R Sibley, Rupert Faraway, Andrea S Elser, Anob M Chakrabarti, Zhen Wang, Julian Koenig, David Perera, Vihandha O Wickramasinghe, Ashok R Venkitaraman, Nicholas M Luscombe, Luciano Saieva, Livio Pellizzoni, Christopher WJ Smith, Tomaz Curk, Jernej Ule

Nature Structural and Molecular Biology | NATURE PUBLISHING GROUP | Published : 2019

Abstract

Studies of spliceosomal interactions are challenging due to their dynamic nature. Here we used spliceosome iCLIP, which immunoprecipitates SmB along with small nuclear ribonucleoprotein particles and auxiliary RNA binding proteins, to map spliceosome engagement with pre-messenger RNAs in human cell lines. This revealed seven peaks of spliceosomal crosslinking around branchpoints (BPs) and splice sites. We identified RNA binding proteins that crosslink to each peak, including known and candidate splicing factors. Moreover, we detected the use of over 40,000 BPs with strong sequence consensus and structural accessibility, which align well to nearby crosslinking peaks. We show how the position ..

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Grants

Awarded by European Research Council


Awarded by Slovenian Research Agency


Awarded by Royal Society


Awarded by Biotechnology and Biological Sciences Research Council


Awarded by Wellcome Trust


Awarded by Medical Research Council


Awarded by National Institute of Neurological Disorders and Stroke of the National Institutes of Health (NIH-NINDS)


Awarded by Cancer Research UK


Awarded by UK Medical Research Council


Awarded by NATIONAL INSTITUTE OF NEUROLOGICAL DISORDERS AND STROKE


Funding Acknowledgements

We thank M. Llorian for help with the in vitro splicing reactions, K. Zarnack and G. Rot for help with the data analyses and L. Strittmatter and members of the Ule lab for helpful discussions and comments on the manuscript. This work was supported primarily by the European Research Council (grant nos. 206726-CLIP and 617837-Translate) and the Slovenian Research Agency (grant nos. P2-0209, Z7-3665 and J7-5460). C.R.S. was supported by an Edmond Lily Safra Fellowship and a Sir Henry Dale Fellowship jointly funded by the Wellcome Trust and the Royal Society (grant no. 215454/Z/19/Z). A.S.E. is supported by the Biotechnology and Biological Sciences Research Council (grant no. BB/M009513/1). A.M.C. is supported by a Wellcome Trust PhD Training Fellowship for Clinicians (no. 110292/Z/15/Z). D. P. and V.O.W. were supported by Medical Research Council grants (nos. MC_UU_12022/1 and MC_UU_12022/8 to A.R.V). L.P. was supported by the National Institute of Neurological Disorders and Stroke of the National Institutes of Health (NIH-NINDS) (grant no. R01 NS102451). The Francis Crick Institute receives its core funding from Cancer Research UK (grant no. FC001002), the UK Medical Research Council (grant no. FC001002) and the Wellcome Trust (grant no. FC001002).