Programmable Recruitment of RNA-Binding Proteins Enables Small Molecule-Directed Destabilization of Nuclear Pre-mRNA

Chemically induced proximity has not been systematically applied to control RNA fate. Here, a programmable platform was developed to identify RNA-binding proteins (RBPs) that can be recruited by small molecules to destabilize RNA. Using microtubule-associated protein Tau (MAPT) pre-mRNA as a model target, a heterobifunctional molecule was designed to bind both a ligandable structure in MAPT pre-mRNA and FKBP12F36V-tagged RBPs. Screening of a library of tagged RBPs identified several proteins that reduced MAPT RNA levels, including zinc finger protein 36 (ZFP36) and nanos C2HC-type zinc finger 3 (NANOS3). The approach was then extended from engineered proteins to an endogenous RBP. Using small molecule ligandability maps, a cysteine-reactive ligand for ZFP36 was identified. When this ligand was linked to the MAPT-binding small molecule, endogenous ZFP36 was recruited to MAPT mRNA, reducing its abundance in cells. Genetic and chemical controls demonstrated that activity was dependent on both RNA binding and ZFP36 recruitment, supporting an induced-proximity mechanism. These studies establish a general strategy for identifying new recruitable RBP effectors and should advance ribonuclease-targeting chimera (RiboTAC) technology by expanding the repertoire of effector proteins that can be harnessed for RNA degradation. More broadly, new effectors can be discovered through model reporter-based screens and translated to endogenous systems by mining known protein binders and ligandability maps, providing a systematic path to develop small molecules that control RNA stability, including RNAs targeted through structured regions of nuclear pre-mRNAs.