Two cysteine residues in the DNA-binding domain of CREB control binding to CRE and CREB-mediated gene expression

Itamar Goren, Einat Tavor, Amiram Goldblum, Alik Honigman*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

38 Scopus citations

Abstract

The cAMP-responsive element-binding protein (CREB) has been implicated in the regulation of numerous physiological functions including those of several hypoxia-responding genes. All CREB transcription-regulated genes harbor the eight base-pair cAMP-responsive element (CRE) or the seven base-pair AP-1 sequence. Utilizing mutational analysis and biochemical assays, we found that reduction of two cysteine residues located in the DNA-binding basic domain of CREB, enhances the binding efficiency of CREB to DNA and regulates CRE-mediated gene expression. Substitution of these residues to serine renders insensitivity to reduction, hypoxia and to the sulfhydryl-specific modifying agent, N-ethylmaleimide. These substitutions enhance the binding of CREB to its cognate DNA sites under oxidative conditions, and of the CREB-dependent gene expression during normoxia. These findings are supported by results of molecular modeling of the CREB-CRE interactions. We also found that HTLV-1 Tax enhancement of CREB binding to the cellular and the viral DNA sites and activation of the CRE-dependent gene expression are independent of CREB activation exerted by redox conditions. The genetic biochemical and molecular modeling presented in this work indicate that the two cysteine residues in the bZIP domain of CREB regulate the binding efficiency of CREB to its cognate DNA sites and as a consequence the activation of CREB-mediated gene expression.

Original languageAmerican English
Pages (from-to)695-709
Number of pages15
JournalJournal of Molecular Biology
Volume313
Issue number4
DOIs
StatePublished - 2 Nov 2001

Bibliographical note

Funding Information:
We are grateful to Dr Marc Montminy at Harvard Medical School for promptly sending us the plasmid construct pRSVCREB. We are indebted to Dr Richard G. Brennan for the coordinates of CREB bZIP/CRE. We thank Dr Amos Panet for critical reading of the manuscript and useful comments. This work was supported, partially, by the Oppenheimer Endowment Fund for Human Health and by the Alex Grass Center for Drug Design and Synthesis at the School of Pharmacy, Hebrew University of Jerusalem.

Keywords

  • CRE
  • CREB
  • Hypoxia
  • Luciferase
  • bZIP

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