Tristetraprolin Inhibits Poly(A)-Tail Synthesis in Nuclear mRNA that Contains AU-Rich Elements by Interacting with Poly(A)-Binding Protein Nuclear 1

Abstract

Background Tristetraprolin binds mRNA AU-rich elements and thereby facilitates the destabilization of mature mRNA in the cytosol. Methodology/Principal Findings To understand how tristetraprolin mechanistically functions, we biopanned with a phage-display library for proteins that interact with tristetraprolin and retrieved, among others, a fragment of poly(A)-binding protein nuclear 1, which assists in the 3'-polyadenylation of mRNA by binding to immature poly(A) tails and thereby increases the activity of poly(A) polymerase, which is directly responsible for polyadenylation. The tristetraprolin/poly(A)-binding protein nuclear 1 interaction was characterized using tristetraprolin and poly(A)-binding protein nuclear 1 deletion mutants in pull-down and co-immunoprecipitation assays. Tristetraprolin interacted with the carboxyl-terminal region of poly(A)-binding protein nuclear 1 via its tandem zinc finger domain and another region. Although tristetraprolin and poly(A)-binding protein nuclear 1 are located in both the cytoplasm and the nucleus, they interacted in vivo in only the nucleus. In vitro , tristetraprolin bound both poly(A)-binding protein nuclear 1 and poly(A) polymerase and thereby inhibited polyadenylation of AU-rich element–containing mRNAs encoding tumor necrosis factor α, GM-CSF, and interleukin-10. A tandem zinc finger domain–deleted tristetraprolin mutant was a less effective inhibitor. Expression of a tristetraprolin mutant restricted to the nucleus resulted in downregulation of an AU-rich element–containing tumor necrosis factor α/luciferase mRNA construct. Conclusion/Significance In addition to its known cytosolic mRNA–degrading function, tristetraprolin inhibits poly(A) tail synthesis by interacting with poly(A)-binding protein nuclear 1 in the nucleus to regulate expression of AU-rich element–containing mRNA.

A Masked PY-NLS in Drosophila TIS11 and Its Mammalian Homolog Tristetraprolin

“…In yeast, Cth2 has been shown to modulate the selection of the poly(A) site of several mRNAs [46]. More recently, studies have shown that TIS11B regulates the 3’ end processing of the Dll4 mRNA [47], and that TTP could play a role in the polyadenylation of ARE-containing mRNAs via its interaction with poly(A)-binding protein nuclear 1 [48]. It will be of interest to determine whether dTIS11 also participates in nuclear processes and to establish how the Transportin/TIS11 interaction is modulated to intervene in the various roles of TIS11 proteins.…”

Determinants and implications of mRNA poly(A) tail size – Does this protein make my tail look big?

“…AREs and GREs have been demonstrated to induce rapid deadenylation and subsequent decay of mRNAs, which may be mediated by recruitment of the exosome [68] or 5′-3′ decay machinery [27]. The two classes of 3′ UTR elements serve as binding sites for a multitude of AU-binding proteins (AUBPs) or GU-binding proteins, which can enhance or prevent mRNA decay, in some cases specifically via interaction with the poly(A) tail [65,69,70]. …”

“…Tristetraprolin (TTP) is notorious for promoting cytoplasmic decay of mRNAs containing AREs, but in the nucleus it can directly interact with PABN1 and PAP to inhibit polyadenylation, potentially contributing to a decrease in nuclear export of mRNAs [70,88]. The Hu proteins, which stabilize mRNAs in the cytoplasm, interact with CstF and CPSF during transcriptional 3′ end processing to prevent polyadenylation at poly(A) sites containing U-rich sequences [89].…”

Nuclear poly(A) tail size is regulated by Cnot1 during the serum response

“…A good candidate for the regulation of the initial poly(A) tails size of serum response mRNAs is Zfp36 (tristetraprolin), a protein that is known to bind the AU rich elements that are common in serum response mRNAs and regulates their stability in the cytoplasm [15]. Zpf36 has also been shown to interact with Pabpn1 and inhibit nuclear polyadenylation [47]. In addition, U rich element binding proteins from the Elavl (Hu) family have been shown to affect the addition of the poly(A) tail in vitro [48] and Actb1 mRNA has extensive binding sites for this protein in its 3' UTR [49].…”