Pseudouridine synthase 7 is an opportunistic enzyme that binds and modifies substrates with diverse sequences and structures

Abstract

Significance Pseudouridine is among the most-abundant RNA modifications. We present a framework for conceptualizing how eukaryotic pseudouridine synthases select their substrates. This work reveals the structure of yeast pseudouridine synthase 7 (Pus7) and presents cell-based and biochemical investigations of enzyme binding and activity. We demonstrate that Pus7 interacts promiscuously with RNAs containing UG U AR sequences. Our observations raise the question of why these enzymes only modify <5% of UG U AR sequences in the transcriptome, suggesting that factors beyond inherent enzyme properties—such as protein localization, local RNA structure, and RNA–protein interactions—principally shape Pus7 substrate selection. These findings support a role for Pus7 in providing cells with a mechanism to rapidly alter protein synthesis in response to cellular conditions.