Structural and functional adaptation of Haloferax volcanii TFEα/β.

Fabian Blombach,Nanruoyi Zhou,Zoja Soloviev,Thorsten Allers,Tanya Prentice,Mark Zhang,Darya Ausiannikava,Angelo Miguel Figueiredo,Finn Werner,Konstantinos Thalassinos

doi:10.1093/nar/gkx1302

Fabian Blombach, Nanruoyi Zhou + Show 8 more

Open Access

https://doi.org/10.1093/nar/gkx1302

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Abstract

The basal transcription factor TFE enhances transcription initiation by catalysing DNA strand-separation, a process that varies with temperature and ionic strength. Canonical TFE forms a heterodimeric complex whose integrity and function critically relies on a cubane iron-sulphur cluster residing in the TFEβ subunit. Halophilic archaea such as Haloferax volcanii have highly divergent putative TFEβ homologues with unknown properties. Here, we demonstrate that Haloferax TFEβ lacks the prototypical iron-sulphur cluster yet still forms a stable complex with TFEα. A second metal cluster contained in the zinc ribbon domain in TFEα is highly degenerate but retains low binding affinity for zinc, which contributes to protein folding and stability. The deletion of the tfeB gene in H. volcanii results in the aberrant expression of approximately one third of all genes, consistent with its function as a basal transcription initiation factor. Interestingly, tfeB deletion particularly affects foreign genes including a prophage region. Our results reveal the loss of metal centres in Hvo transcription factors, and confirm the dual function of TFE as basal factor and regulator of transcription.

Highlights

The RNA polymerase (RNAP), auxiliary factors and molecular mechanisms of transcription are conserved between archaea and eukaryotes
We constructed the corresponding winged helix-turnedhelix (WH) deletion variant of Haloferax volcanii (Hvo) TFE␤ and verified its complex formation with TFE␣, which confirms that the WH domain is dispensable for dimer formation in Hvo and demonstrates that the domain requirements for dimerization are conserved between Sulfolobus solfataricus (Sso) and Hvo TFE␣/␤ (Figure 2B)
The magnitude of TFE stimulation varies with the promoter context, and since the steady state levels of TFE␤ in Sso rapidly decrease in response to oxidative stress and upon entering stationary phase the genes controlled by promoters that respond stronger to TFE will be down regulated significantly more compared to genes under the control of relatively unresponsive promoters [2]

Summary

Introduction

The RNA polymerase (RNAP), auxiliary factors and molecular mechanisms of transcription are conserved between archaea and eukaryotes. The molecular engineering challenges of initiating transcription in a start site-specific fashion are independent of the type of RNAP and the domain of life. RNAPs need to engage with the promoter region of the double-stranded DNA template, the two DNA strands need to be separated, the template strand loaded into the RNAP active site, and individual NTP substrates bound in a geometry conducive to catalysis. All cellular RNAPs depend on additional basal transcription factors to facilitate this process. In archaea three basal transcription factors (TBP, TFB and TFE) assist RNAP during initiation. TBP and TFB bind to the TATA and BRE motifs and are necessary and sufficient for RNA polymerase recruitment to the promoter and transcription initiation. TFE is not strictly required for transcription in vitro, but stimulates transcription initiation by enhancing open complex formation (OC), i.e. DNA melting and template loading [1]

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