mRNA Openers and Closers: Modulating AU-Rich Element-Controlled mRNA Stability by a Molecular Switch in mRNA Secondary Structure

• Published in: Chembiochem : a European journal of chemical biology 2004-10, Vol.5 (10), p.1432-1447
• Main Authors: Meisner, Nicole-Claudia, Hackermüller, Jörg, Uhl, Volker, Aszódi, András, Jaritz, Markus, Auer, Manfred
• Format: Article
• Language: English
• Subjects: Antigens, Surface - genetics; Antigens, Surface - metabolism; AU-rich elements; Base Pairing; Base Sequence; Binding Sites - physiology; ELAV Proteins; ELAV-Like Protein 1; functional RNAs; Humans; HuR; Molecular Sequence Data; Nucleic Acid Conformation; RNA recognition; RNA Stability - genetics; RNA Stability - physiology; RNA structures; RNA, Messenger - chemistry; RNA, Messenger - genetics; RNA, Messenger - metabolism; RNA-Binding Proteins - genetics; RNA-Binding Proteins - metabolism; Structure-Activity Relationship; systems biology; Time Factors
• ISSN: 1439-4227; 1439-7633; 1439-4227
• DOI: 10.1002/cbic.200400219

Approximately 3 000 genes are regulated in a time-, tissue-, and stimulus-dependent manner by degradation or stabilization of their mRNAs. The process is mediated by interaction of AU-rich elements (AREs) in the mRNA's 3′-untranslated regions with trans-acting factors. AU-rich element-controlled genes of fundamentally different functional relevance depend for their activation on one positive regulator, HuR. Here we present a methodology to exploit this central regulatory process for specific manipulation of AU-rich element-controlled gene expression at the mRNA level. With a combination of single-molecule spectroscopy, computational biology, and molecular and cellular biochemistry, we show that mRNA recognition by HuR is dependent on the presentation of the sequence motif NNUUNNUUU in single-stranded conformation. The presentation of the HuR binding site in the mRNA secondary structure appears to act analogously to a regulatory on/off switch that specifically controls HuR access to mRNAs in cis. Based on this finding we present a methodology for manipulating ARE mRNA levels by actuating this conformational switch specifically in a target mRNA. Computationally designed oligonucleotides (openers) enhance the NNUUNNUUU accessibility by rearranging the mRNA conformation. Thereby they increase in vitro and endogenous HuR-mRNA complex formation which leads to specific mRNA stabilization (as demonstrated for TNFα and IL-2, respectively). Induced HuR binding both inside and outside the AU-rich element promotes functional IL-2 mRNA stabilization. This opener-induced mRNA stabilization mimics the endogenous IL-2 response to CD28 stimulation in human primary T-cells. We therefore propose that controlled modulation of the AU-rich element conformation by mRNA openers or closers allows message stabilization or destabilization in cis to be specifically triggered. The described methodology might provide a means for studying distinct pathways in a complex cellular network at the node of mRNA stability control. It allows ARE gene expression to be potentially silenced or boosted. This will be of particular value for drug-target validation, allowing the diseased phenotype to ameliorate or deteriorate. Finally, the mRNA openers provide a rational starting point for target-specific mRNA stability assays to screen for low-molecular-weight compounds acting as inhibitors or activators of an mRNA structure rearrangement.