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Series GSE156133 Query DataSets for GSE156133
Status Public on Oct 27, 2020
Title Widespread transcriptional readthrough caused by Nab2 depletion leads to chimeric transcripts with retained introns
Organism Saccharomyces cerevisiae
Experiment type Other
Summary Nascent RNA sequencing has recently revealed that pre-mRNA splicing can occur shortly after the intron emerges from RNA polymerase II (Pol II). Differences in co-transcriptional splicing profiles suggest regulation by cis- and/or trans-acting factors. Here we used Single Molecule Intron Tracking (SMIT) in budding yeast to identify a cohort of regulators by machine learning. One candidate, Nab2, displayed reduced co-transcriptional splicing of some pre-mRNAs when depleted. Unexpectedly, these splicing defects were attributable to readthrough transcription, which was revealed by long read sequencing of nascent RNA; individual readthrough transcripts induced by Nab2 depletion sometimes spanned multiple genes. Thus, Nab2 regulation of splicing was indirect. Moreover, unspliced transcripts displayed downstream readthrough in both control and Nab2-depleted cells, highlighting the coupling between splicing and 3′ end formation. We conclude that Nab2 is required for proper 3′ end processing, which ensures both transcription termination as well as proper splicing of downstream genes.
Overall design Single Molecule Intron Tracking (SMIT) was performed for 2 replicates each of 6 different yeast deletion strains as well as the wild type control. Additional SMIT libraries were prepared for Anchor-Away strains (Nab2-AA and Control-AA). Finally, long read sequencing libraries were prepared from nascent RNA extracted from the Nab2-AA and Control-AA samples in duplicate.

Please note that the 'Nab2AA_ControlAA_fast5.tar.gz' (linked to the Long read Nab2-AA rep1 sample records) contains raw data for all four "Long read" samples. As the algorithm for converting and demultiplexing the fast5 into fastq files is proprietary software and not all users will have access to the exact form/version used, both fastq and fast5 data were made available as SRA and GEO records, respectively.
Contributor(s) Alpert T, Straube K, Carrillo Oesterreich F, Neugebauer KM
Citation(s) 33113357
Submission date Aug 12, 2020
Last update date Jan 04, 2021
Contact name Tara Diane Alpert
Organization name Yale University
Department Molecular Biophysics and Biochemistry
Lab Karla Neugebauer
Street address 333 Cedar St.
City New Haven
State/province CT
ZIP/Postal code 06511
Country USA
Platforms (2)
GPL17342 Illumina HiSeq 2500 (Saccharomyces cerevisiae)
GPL25739 MinION (Saccharomyces cerevisiae)
Samples (24)
GSM4725167 SMIT WT rep1
GSM4725168 SMIT WT rep2
GSM4725169 SMIT Npl3-KO rep1
BioProject PRJNA656824
SRA SRP277240

Download family Format
SOFT formatted family file(s) SOFTHelp
MINiML formatted family file(s) MINiMLHelp
Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE156133_RAW.tar 193.8 Gb (http)(custom) TAR (of BED, TAR, TXT)
GSE156133_dSMIT_deletions_rep1.txt.gz 3.2 Kb (ftp)(http) TXT
GSE156133_dSMIT_deletions_rep2.txt.gz 3.3 Kb (ftp)(http) TXT
GSE156133_longread_fractionspliced.txt.gz 4.7 Kb (ftp)(http) TXT
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file
Processed data are available on Series record

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