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Schizosaccharomyces pombe Grx4 regulates the transcriptional repressor Php4 via [2Fe-2S] cluster binding

Adrienne C Dlouhy1, Jude Beaudoin, Simon Labbé

  • 1Department of Chemistry and Biochemistry, University of South Carolina, 631 Sumter Street, Columbia, South Carolina 29208, USA. outten@mailbox.sc.edu.

Metallomics : Integrated Biometal Science
|July 21, 2017
PubMed

Insights

Fission yeast protein Grx4 controls iron regulation by binding a bridging [2Fe-2S] cluster, impacting the transcription factor Php4. This biochemical study reveals essential cysteine residues for Fe-S cluster binding and complex stability.

Area of Science:

  • Molecular Biology
  • Biochemistry
  • Yeast Genetics

Background:

  • Fission yeast *Schizosaccharomyces pombe* regulates iron usage via the CCAAT-binding factor Php4.
  • Php4 function and localization are modulated by cytosolic glutaredoxin Grx4 in an iron-dependent manner.
  • Understanding the precise biochemical mechanisms of Grx4-Php4 interaction is crucial for iron homeostasis.

Purpose of the Study:

  • To biochemically characterize the protein-protein and protein-metal interactions between Grx4 and Php4.
  • To elucidate the role of iron-sulfur clusters in the regulation of Php4 by Grx4.
  • To identify key residues involved in these interactions.

Main Methods:

  • Spectroscopic analysis of [2Fe-2S] cluster binding to Grx4.
  • Copurification and spectroscopic characterization of the Grx4-Php4 complex.
  • In vitro titration experiments to assess complex formation and interconvertibility.
  • Site-directed mutagenesis of conserved cysteines in Grx4 and Php4.

Main Results:

  • Grx4 binds a [2Fe-2S] cluster with characteristic spectroscopic features.
  • Grx4 and Php4 form a complex containing a distinct [2Fe-2S] cluster.
  • Conserved cysteines (Grx4 Cys172, Php4 Cys221/Cys227) are essential for Fe-S cluster binding and complex stability.
  • Fe-S cluster interconversion appears limited without additional factors.

Conclusions:

  • Grx4 regulates Php4 activity through the binding of a bridging [2Fe-2S] cluster.
  • This mechanism highlights the role of iron-sulfur clusters in gene expression regulation.
  • Conserved cysteines are critical for the structural integrity and function of the Grx4-Php4 regulatory complex.

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