Activation by NarL at the Escherichia coli ogt promoter

TY - JOUR

T1 - Activation by NarL at the Escherichia coli ogt promoter

AU - Ruanto, Patcharawarin

AU - Chismon, David J

AU - Hothersall, Joanne

AU - Godfrey, Rita E

AU - Lee, David J

AU - Busby, Stephen J W

AU - Browning, Douglas F

N1 - © 2020 The Author(s).

PY - 2020/8/14

Y1 - 2020/8/14

N2 - The Escherichia coli NarX/NarL two-component response regulator system regulates gene expression in response to nitrate ions and the NarL protein is a global transcription factor, which activates transcript initiation at many target promoters. One such target, the E. coliogt promoter, which controls the expression of an O6‑alkylguanine-DNA-alkyltransferase, is dependent on NarL binding to two DNA targets centred at positions -44.5 and -77.5 upstream from the transcript start. Here, we describe ogt promoter derivatives that can be activated solely by NarL binding either at position -44.5 or position -77.5. We show that NarL can also activate the ogt promoter when located at position -67.5. We present data to argue that NarL-dependent activation of transcript initiation at the ogt promoter results from a direct interaction between NarL and a determinant in the C-terminal domain of the RNA polymerase α subunit. Footprinting experiments show that, at the -44.5 promoter, NarL and the C-terminal domain of the RNA polymerase α subunit bind to opposite faces of promoter DNA, suggesting an unusual mechanism of transcription activation. Our work suggests new organisations for activator-dependent transcription at promoters and future applications for biotechnology.

AB - The Escherichia coli NarX/NarL two-component response regulator system regulates gene expression in response to nitrate ions and the NarL protein is a global transcription factor, which activates transcript initiation at many target promoters. One such target, the E. coliogt promoter, which controls the expression of an O6‑alkylguanine-DNA-alkyltransferase, is dependent on NarL binding to two DNA targets centred at positions -44.5 and -77.5 upstream from the transcript start. Here, we describe ogt promoter derivatives that can be activated solely by NarL binding either at position -44.5 or position -77.5. We show that NarL can also activate the ogt promoter when located at position -67.5. We present data to argue that NarL-dependent activation of transcript initiation at the ogt promoter results from a direct interaction between NarL and a determinant in the C-terminal domain of the RNA polymerase α subunit. Footprinting experiments show that, at the -44.5 promoter, NarL and the C-terminal domain of the RNA polymerase α subunit bind to opposite faces of promoter DNA, suggesting an unusual mechanism of transcription activation. Our work suggests new organisations for activator-dependent transcription at promoters and future applications for biotechnology.

KW - molecular interactions

KW - NarL, promoters

KW - transcription activation

UR - http://www.scopus.com/inward/record.url?scp=85089300915&partnerID=8YFLogxK

U2 - 10.1042/BCJ20200408

DO - 10.1042/BCJ20200408

M3 - Article

C2 - 32662815

VL - 477

SP - 2807

EP - 2820

JO - Biochem J

JF - Biochem J

SN - 0264-6021

IS - 15

ER -