Promoter-sequence determinants and structural basis of primer-dependent transcription initiation in Escherichia coli

Skalenko, Kyle, Li, Lingting, Zhang, Yuanchao, Vvedenskaya, Irina, Winkelman, Jared, Cope, Alexander, Taylor, Deanne, Shah, Premal, Ebright, Richard, Kinney, Justin, Zhang, Yu, Nickels, Bryce (April 2021) Promoter-sequence determinants and structural basis of primer-dependent transcription initiation in Escherichia coli. BioRxiv. (Unpublished)

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Abstract

Chemical modifications of RNA 5′ ends enable “epitranscriptomic” regulation, influencing multiple aspects of RNA fate. In transcription initiation, a large inventory of substrates compete with nucleoside triphosphates (NTPs) for use as initiating entities, providing an ab initio mechanism for altering the RNA 5′ end. In Escherichia coli cells, RNAs with a 5′-end hydroxyl are generated by use of dinucleotide RNAs as primers for transcription initiation, “primer-dependent initiation.” Here we use massively systematic transcript end readout (“MASTER”) to detect and quantify RNA 5′ ends generated by primer-dependent initiation for ~410 (~1,000,000) promoter sequences in E. coli. The results show primer-dependent initiation in E. coli involves any of the 16 possible dinucleotide primers and depends on promoter sequences in, upstream, and downstream of the primer binding site. The results yield a consensus sequence for primer-dependent initiation, YTSS-2NTSS-1NTSSWTSS+1, where TSS is the transcription start site, NTSS-1NTSS is the primer binding site, Y is pyrimidine, and W is A or T. Biochemical and structure-determination studies show that the base pair (nontemplate-strand base:template-strand base) immediately upstream of the primer binding site (Y:RTSS-2, where R is purine) exerts its effect through the base on the DNA template strand (RTSS-2) through inter-chain base stacking with the RNA primer. Results from analysis of a large set of natural, chromosomally-encoded E. coli promoters support the conclusions from MASTER. Our findings provide a mechanistic and structural description of how TSS-region sequence hard-codes not only the TSS position, but also the potential for epitranscriptomic regulation through primer-dependent transcription initiation.

Item Type:	Paper
Subjects:	bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > transcription organism description > bacteria > escherichia coli
CSHL Authors:	Kinney, Justin B.
Communities:	CSHL labs > Kinney lab
SWORD Depositor:	CSHL Elements
Depositing User:	CSHL Elements
Date:	7 April 2021
Date Deposited:	10 May 2022 14:31
Last Modified:	20 May 2024 19:46
URI:	https://repository.cshl.edu/id/eprint/40606

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