Ssl2/TFIIH function in transcription start site scanning by RNA Polymerase II in Saccharomyces cerevisiae
- University of Pittsburgh, United States
- Rutgers University, United States
- Cornell University, United States
Abstract
In Saccharomyces cerevisiae, RNA Polymerase II (Pol II) selects transcription start sites (TSS) by a unidirectional scanning process. During scanning, a preinitiation complex (PIC) assembled at an upstream core promoter initiates at select positions within a window ~40-120 basepairs downstream. Several lines of evidence indicate that Ssl2, the yeast homolog of XPB and an essential and conserved subunit of the general transcription factor (GTF) TFIIH, drives scanning through its DNA-dependent ATPase activity, therefore potentially controlling both scanning rate and scanning extent (processivity). To address questions of how Ssl2 functions in promoter scanning and interacts with other initiation activities, we leveraged distinct initiation-sensitive reporters to identify novel ssl2 alleles. These ssl2 alleles, many of which alter residues conserved from yeast to human, confer either upstream or downstream TSS shifts at the model promoter ADH1 and genome-wide. Specifically, tested ssl2 alleles alter TSS selection by increasing or narrowing the distribution of TSSs used at individual promoters. Genetic interactions of ssl2 alleles with other initiation factors are consistent with ssl2 allele classes functioning through increasing or decreasing scanning processivity but not necessarily scanning rate. These alleles underpin a residue interaction network that likely modulates Ssl2 activity and TFIIH function in promoter scanning. We propose that the outcome of promoter scanning is determined by two functional networks, the first being Pol II activity and factors that modulate it to determine initiation efficiency within a scanning window, and the second being Ssl2/TFIIH and factors that modulate scanning processivity to determine the width of the scanning widow.
Data availability
Genomics datasets generated in the current study are available in the NCBI BioProject and SRA, under the accession numbers of PRJNA681384 and SRP295731, respectively. The processed genomic data files are available in GEO, under the accession number of GSE182792. The streamlined commands to generate TSS-seq bedGraph files, count tables, tables of expression, spread and median TSS can be found at https://github.com/Kaplan-Lab-Pitt/Ssl2_scanning.
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Ssl2/TFIIH Function in Transcription Start Site Scanning by RNA Polymerase II in Saccharomyces cerevisiaeNCBI Gene Expression Omnibus, GSE182792.
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Function of Ssl2/TFIIH in RNA Polymerase II Transcription Start Site ScanningNCBI Gene Expression Omnibus, SRP295731.
Article and author information
Author details
Funding
National Institute of General Medical Sciences (R01GM120450)
- Craig D Kaplan
National Institute of General Medical Sciences (R01GM059055)
- B Franklin Pugh
National Institute of General Medical Sciences (R01GM059055)
- Bryce E Nickels
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2021, Zhao et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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