The debranching enzyme Dbr1 regulates lariat turnover and intron splicing

Luke Buerer, Nathaniel E. Clark, Anastasia Welch, Chaorui Duan, Allison J. Taggart, Brittany A. Townley, Jing Wang, Rachel Soemedi, Stephen Rong, Chien-Ling Lin, Yi Zeng, Adam Katolik, Jonathan P. Staley, Masad J. Damha, Nima Mosammaparast and William G. Fairbrother ()
Additional contact information
Luke Buerer: Brown University
Nathaniel E. Clark: Brown University
Anastasia Welch: Brown University
Chaorui Duan: Brown University
Allison J. Taggart: Brown University
Brittany A. Townley: Washington University School of Medicine
Jing Wang: Brown University
Rachel Soemedi: Brown University
Stephen Rong: Brown University
Chien-Ling Lin: Brown University
Yi Zeng: University of Chicago
Adam Katolik: McGill University
Jonathan P. Staley: University of Chicago
Masad J. Damha: McGill University
Nima Mosammaparast: Washington University School of Medicine
William G. Fairbrother: Brown University

Nature Communications, 2024, vol. 15, issue 1, 1-13

Abstract: Abstract The majority of genic transcription is intronic. Introns are removed by splicing as branched lariat RNAs which require rapid recycling. The branch site is recognized during splicing catalysis and later debranched by Dbr1 in the rate-limiting step of lariat turnover. Through generation of a viable DBR1 knockout cell line, we find the predominantly nuclear Dbr1 enzyme to encode the sole debranching activity in human cells. Dbr1 preferentially debranches substrates that contain canonical U2 binding motifs, suggesting that branchsites discovered through sequencing do not necessarily represent those favored by the spliceosome. We find that Dbr1 also exhibits specificity for particular 5’ splice site sequences. We identify Dbr1 interactors through co-immunoprecipitation mass spectrometry. We present a mechanistic model for Dbr1 recruitment to the branchpoint through the intron-binding protein AQR. In addition to a 20-fold increase in lariats, Dbr1 depletion increases exon skipping. Using ADAR fusions to timestamp lariats, we demonstrate a defect in spliceosome recycling. In the absence of Dbr1, spliceosomal components remain associated with the lariat for a longer period of time. As splicing is co-transcriptional, slower recycling increases the likelihood that downstream exons will be available for exon skipping.

Date: 2024
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DOI: 10.1038/s41467-024-48696-1

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