Ligand binding reveals a role for heme in translationally-controlled tumor protein dimerization.

Andrew T Lucas,Xiangping Fu,Daniel G S Capelluto,Jingjing Liu,Carla V Finkielstein,Jianhua Yang,Mary K Brannon,Vladimir N Uversky

doi:10.1371/journal.pone.0112823

Abstract

The translationally-controlled tumor protein (TCTP) is a highly conserved, ubiquitously expressed, abundant protein that is broadly distributed among eukaryotes. Its biological function spans numerous cellular processes ranging from regulation of the cell cycle and microtubule stabilization to cell growth, transformation, and death processes. In this work, we propose a new function for TCTP as a “buffer protein” controlling cellular homeostasis. We demonstrate that binding of hemin to TCTP is mediated by a conserved His-containing motif (His76His77) followed by dimerization, an event that involves ligand-mediated conformational changes and that is necessary to trigger TCTP's cytokine-like activity. Mutation in both His residues to Ala prevents hemin from binding and abrogates oligomerization, suggesting that the ligand site localizes at the interface of the oligomer. Unlike heme, binding of Ca2+ ligand to TCTP does not alter its monomeric state; although, Ca2+ is able to destabilize an existing TCTP dimer created by hemin addition. In agreement with TCTP's proposed buffer function, ligand binding occurs at high concentration, allowing the “buffer” condition to be dissociated from TCTP's role as a component of signal transduction mechanisms.

Highlights

The translationally-controlled tumor protein (TCTP) is a highly conserved and ubiquitously expressed eukaryotic protein whose cellular function spans from mechanisms of cell growth and division to cytoskeleton reorganization and cell morphology
In an attempt to consolidate some of the conflicting data surrounding the role of ligand binding in TCTP function, its influence in TCTP behavior, as well as the nature of their biochemical interaction in a comprehensive model, we evaluated the role of known interactors, such as heme and Ca2+, in modulating structural rearrangements associated with TCTP oligomerization and ligand binding
Ligand binding influences the oligomeric state of TCTP Recombinant TCTP was expressed in E. coli as a glutathione S-transferase (GST) N

Summary

Introduction

The translationally-controlled tumor protein (TCTP) is a highly conserved and ubiquitously expressed eukaryotic protein whose cellular function spans from mechanisms of cell growth and division to cytoskeleton reorganization and cell morphology (for review see [1]). Other evidence establishes a role for TCTP in cell proliferation This includes i) the regulation of the GTPase activity of the Drosophila Ras homologue Rheb, a direct target of TSC1/2 tumor suppressors responsible for tuberous sclerosis [8], ii) the stabilization of the GDP form of the translational elongation factor eEF1A [11], and iii) progression through cytokinesis by a mechanism that involves phosphorylation of TCTP by the polo-like kinase and, reduction of the microtubule-stabilizing activity of TCTP [12],[13]. Unlike its mononuclear cellular activated version, the serum form of human HRF (similar to extracellular TCTP) exhibits cytokine-like activity in vivo when dimerized, an event that is independent of post-translational modifications and thought to be mediated by a largely unknown player(s) [20]

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Conclusion