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Cell. 2018 May 3;173(4):1014-1030.e17. doi: 10.1016/j.cell.2018.03.020.

Spliceosome Profiling Visualizes Operations of a Dynamic RNP at Nucleotide Resolution.

Author information

1
Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA 94158, USA.
2
Department of Statistics, University of California, Los Angeles, Los Angeles, CA, USA.
3
Department of Molecular Medicine, the Scripps Research Institute, La Jolla, CA, USA.
4
Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA 94158, USA; Chan-Zuckerberg Biohub, San Francisco, CA 94158, USA. Electronic address: hitenmadhani@gmail.com.

Abstract

Tools to understand how the spliceosome functions in vivo have lagged behind advances in the structural biology of the spliceosome. Here, methods are described to globally profile spliceosome-bound pre-mRNA, intermediates, and spliced mRNA at nucleotide resolution. These tools are applied to three yeast species that span 600 million years of evolution. The sensitivity of the approach enables the detection of canonical and non-canonical events, including interrupted, recursive, and nested splicing. This application of statistical modeling uncovers independent roles for the size and position of the intron and the number of introns per transcript in substrate progression through the two catalytic stages. These include species-specific inputs suggestive of spliceosome-transcriptome coevolution. Further investigations reveal the ATP-dependent discard of numerous endogenous substrates after spliceosome assembly in vivo and connect this discard to intron retention, a form of splicing regulation. Spliceosome profiling is a quantitative, generalizable global technology used to investigate an RNP central to eukaryotic gene expression.

KEYWORDS:

pre-mRNA splicing; spliceosome; splicing catalysis; splicing fidelity

Comment in

PMID:
29727661
PMCID:
PMC5940017
DOI:
10.1016/j.cell.2018.03.020
[Indexed for MEDLINE]
Free PMC Article

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