High-throughput discovery of regulatory effector domains in human RNA-binding proteins

RNA regulation plays an integral role in tuning gene expression and is controlled by thousands of RNA-binding proteins (RBPs). We develop and use a high-throughput recruitment assay (HT-RNA-Recruit) to identify regulatory domains within human RBPs by recruiting over 30,000 protein tiles from 367 RBPs to a reporter mRNA. We discover over 100 unique RNA-regulatory effectors in 86 distinct RBPs, presenting evidence that RBPs contain functionally separable domains that dictate their post-transcriptional control of gene expression, and identify some with unique activity at 5 or 3UTRs. We identify some domains that downregulate gene expression both when recruited to DNA and RNA, and dissect their mechanisms of regulation. Finally, we build a synthetic RNA regulator that can stably maintain gene expression at desired levels that are predictable by a mathematical model. This work serves as a resource for human RNA-regulatory effectors and expands the synthetic repertoire of RNA-based genetic control tools. Highlights- HT-RNA-Recruit identifies hundreds of RNA-regulatory effectors in human proteins. - Recruitment to 5 and 3 UTRs identifies regulatory domains unique to each position. - Some protein domains have both transcriptional and post-transcriptional regulatory activity. - We develop a synthetic RNA regulator and a mathematical model to describe its behavior.