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prp8 mutations that cause human retinitis pigmentosa lead to a U5 snRNP maturation defect in yeast

Abstract

Prp8 protein (Prp8p) is a highly conserved pre-mRNA splicing factor and a component of spliceosomal U5 small nuclear ribonucleoproteins (snRNPs). Although it is ubiquitously expressed, mutations in the C terminus of human Prp8p cause the retina-specific disease retinitis pigmentosa (RP). The biogenesis of U5 snRNPs is poorly characterized. We present evidence for a cytoplasmic precursor U5 snRNP in yeast that lacks the mature U5 snRNP component Brr2p and depends on a nuclear localization signal in Prp8p for its efficient nuclear import. The association of Brr2p with the U5 snRNP occurs within the nucleus. RP mutations in Prp8p in yeast result in nuclear accumulation of the precursor U5 snRNP, apparently as a consequence of disrupting the interaction of Prp8p with Brr2p. We therefore propose a novel assembly pathway for U5 snRNP complexes that is disrupted by mutations that cause human RP.

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Figure 1: Functional analysis of a nuclear localization signal in Prp8p.
Figure 2: More Prp8p–Aar2p–U5 snRNP complex forms with ΔNLS-Prp8p.
Figure 3: Analysis of Brr2p interactions.
Figure 4: Effects of prp8-rp mutations.
Figure 5: Cartoon showing the proposed pathway for U5 snRNP biogenesis in budding yeast.

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Acknowledgements

We thank P. Fabrizio (Max Planck Institute, Göttingen) for the very generous gift of anti-Aar2p and anti-Snu114p antibodies, O. Cordin (University of Edinburgh) for anti-Brr2p anti-peptide antibodies, M. Spiller for assistance with immunofluorescence staining and microscopy, A. Kutach and C. Guthrie for sharing information before publication, and I. Davis, M. Reijns and D. Tollervey for critical comments. This work was funded by studentships from The Darwin Trust of Edinburgh to K.-L.B. and T.A. and by Wellcome Trust Grants 067311 (J.D. Beggs) and 073988 (C.F.I.) and European Commission grant LSH-2004-518238 (EURASNET Network of Excellence). J.D. Beggs is the Royal Society Darwin Trust Professor.

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K.-L.B. contributed Figures 1, 2, 4c, 4e–h and 5 and Supplementary Figures 1, 3 and 6; R.J.G. contributed Figures 3b, 3c and 4b and Supplementary Figure 2; P.E. contributed Figure 4d and investigated the effects of HA-tagging Prp8p with and without the rp mutations (data not shown); J.D. Barrass performed the RT-PCR and microarray analyses (Supplementary Figs. 4 and 5); T.A. contributed Figure 3a; C.F.I. provided expertise on RP; J.D. Beggs supervised all the work and wrote most of the paper.

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Correspondence to Jean D Beggs.

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Boon, KL., Grainger, R., Ehsani, P. et al. prp8 mutations that cause human retinitis pigmentosa lead to a U5 snRNP maturation defect in yeast. Nat Struct Mol Biol 14, 1077–1083 (2007). https://doi.org/10.1038/nsmb1303

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