A Mechanism for microRNA Arm Switching Regulated by Uridylation

Abstract

Strand selection is a critical step in microRNA (miRNA) biogenesis.Although the dominant strand may alter depending on cellular contexts, the molecular mechanism and physiological significance of such alternative strand selection (or "arm switching") remain elusive. Here we find mir-324 as one of the strongly regulated miRNAs by arm switching, and identify terminal uridylyl transferases TUT4 and TUT7 as the key regulators. Uridylation of pre-mir-324 by TUT4/7 re-positions DICER on the pre-miRNA and shifts the cleavage site. This alternative processing produces a duplex with a different terminus, from which the 3 ′ strand (3p) is selected instead of the 5 ′ strand (5p). In glioblastoma, the TUT4/7 and 3p levels are upregulated while the 5p level is reduced. Manipulation of the strand ratio is sufficient to impair glioblastoma cell proliferation. This study uncovers a role of uridylation as a molecular switch in alternative strand selection and implicates its therapeutic potential.

microRNA strand selection: Unwinding the rules

“…C. elegans miR‐51 isomiRs are shown as an example. (c) Contribution of uridylation to strand selection of miR‐324 (Kim et al, 2020). In wild‐type tissues expressing low levels of terminal 3′ uridylases (TUTases), nonuridylated miR‐324‐5p is selected for miRISC loading.…”

“…One major example is the nontemplated addition of uracil to the 3′ end of a miRNA strand that is catalyzed by 3′ terminal uridyl transferase (TUT) enzymes (reviewed in Menezes, Balzeau, & Hagan, 2018; Pirouz, Ebrahimi, & Gregory, 2019). TUT4 and TUT7, which uridylate let‐7 in an evolutionarily conserved manner (Chang, Triboulet, Thornton, & Gregory, 2013; Hagan, Piskounova, & Gregory, 2009; Heo et al, 2009; Lehrbach et al, 2009; Thornton, Chang, Piskounova, & Gregory, 2012; Wynsberghe et al, 2011), have recently been shown to affect strand selection of human miR‐324 (Kim et al, 2020). TUT4/TUT7 uridylate the 3′ end of miR‐324‐3p, which changes the site of Dicer processing to generate a shorter miR‐324 duplex that undergoes reversed strand selection, stabilizing the miR‐324‐3p strand instead of the typically more abundant miR‐324‐5p strand (Figure 4c; Kim et al, 2020).…”

“…TUT4 and TUT7, which uridylate let‐7 in an evolutionarily conserved manner (Chang, Triboulet, Thornton, & Gregory, 2013; Hagan, Piskounova, & Gregory, 2009; Heo et al, 2009; Lehrbach et al, 2009; Thornton, Chang, Piskounova, & Gregory, 2012; Wynsberghe et al, 2011), have recently been shown to affect strand selection of human miR‐324 (Kim et al, 2020). TUT4/TUT7 uridylate the 3′ end of miR‐324‐3p, which changes the site of Dicer processing to generate a shorter miR‐324 duplex that undergoes reversed strand selection, stabilizing the miR‐324‐3p strand instead of the typically more abundant miR‐324‐5p strand (Figure 4c; Kim et al, 2020). Interestingly, TUT4/TUT7 expression levels appear to be regulated in developmental and tissue‐specific fashions, suggesting that regulation of TUT4/TUT7 activity can dictate strand selection of miR‐324 in humans, resulting in arm switching across tissues (Kim et al, 2020).…”

“…TUT4/TUT7 uridylate the 3′ end of miR‐324‐3p, which changes the site of Dicer processing to generate a shorter miR‐324 duplex that undergoes reversed strand selection, stabilizing the miR‐324‐3p strand instead of the typically more abundant miR‐324‐5p strand (Figure 4c; Kim et al, 2020). Interestingly, TUT4/TUT7 expression levels appear to be regulated in developmental and tissue‐specific fashions, suggesting that regulation of TUT4/TUT7 activity can dictate strand selection of miR‐324 in humans, resulting in arm switching across tissues (Kim et al, 2020).…”

Screening by deep sequencing reveals mediators of microRNA tailing inC. elegans

“…The type of uridylation and its consequence depends on the length of the 3’ overhang, where 1-nt overhangs are mono-uridylated to favor biogenesis and further recessed ends are oligo-uridylated resulting in decay of the precursor (12). Precursor uridylation can also regulate biogenesis by shifting the site of Dicer cleavage, which can result in alternative miRNA guide strand selection (arm-switching) (13). Oligo-uridylation and decay of the let- 7 precursor is induced even in the context of a canonical 2-nt 3’ overhang by binding of the RNA-binding protein LIN-28 (14, 15).…”