Pb(II) coordination to the nonclassical zinc finger tristetraprolin: retained function with an altered fold.

Abstract

Tristetraprolin (TTP) is a nonclassical CCCH zinc finger (ZF) that plays a crucial role in regulating inflammation. TTP regulates cytokine mRNAs by specific binding of its two conserved ZF domains (CysX8CysX5CysX3His) to adenylate-uridylate-rich sequences (AREs) at the 3'-untranslated region, leading to degradation of the RNA. Dysregulation of TTP in animal models has demonstrated several cytokine-related syndromes, including chronic inflammation and autoimmune disorders. Exposure to Pb(II), a prevalent environmental toxin, is known to contribute to similar pathologies, in part by disruption of and/or competition with cysteine-rich metalloproteins. TTP's role during stress as a ubiquitous translational regulator of cell signaling (and dysfunction), which may underpin various phenotypes of Pb(II) toxicity, highlights the importance of understanding the interaction between TTP and Pb(II). The impact of Pb(II) binding on TTP's fold and RNA-binding function was analyzed via UV-Vis spectroscopy, circular dichroism, X-ray absorption spectroscopy, nuclear magnetic resonance spectroscopy, and fluorescence anisotropy. A construct containing the two ZF domains of TTP (TTP-2D) bound to Pb(II) with nanomolar affinity and exhibited a different geometry and fold in comparison to Zn2-TTP-2D. Despite the altered secondary structure, Pb(II)-substituted TTP-2D bound a canonical ARE sequence more selectively than Zn2-TTP-2D. Taken together, these data suggest that Pb(II) may interfere with proper TTP regulation and hinder the cell's ability to respond to inflammation.