Single-Molecule View on the Duality of Microrna Uridylation

Abstract

Terminal uridylyl transferases are a class of enzymes that play a key role in both biogenesis and degradation of microRNAs (miRNAs) in eukaryotes. Previously, we and other groups showed that TUT4 uridylates precursor microRNA (pre-miRNA) in coordination with Lin28 and thus acts as a posttranscriptional repressor of microRNA maturation. Using single-molecule FRET, we show that TUT4 maintains the tight contact with pre-miRNA and Lin28 while it captures the 3' end of RNA and brings this to its catalytic domain. This mechanism leads to the formation of a unique closed loop of the U tail. Besides this repression pathway, terminal uridylyl transferases are also able to enhance miRNA biogenesis through mono-uridylation of pre-miRNA substrates when Lin28 is absent. Using single-molecule fluorescence spectroscopy, we show that TUT7 exhibits preference for group II pre-miRNAs by regulating their binding frequency. In conclusion, our study provides insight into the duality of miRNA uridylation and may give a hint to a general mechanism of action of terminal uridylyl transferases.