TUT4 in Concert with Lin28 Suppresses MicroRNA Biogenesis through Pre-MicroRNA Uridylation

Abstract

As key regulators in cellular functions, microRNAs (miRNAs) themselves need to be tightly controlled. Lin28, a pluripotency factor, was reported to downregulate let-7 miRNA by inducing uridylation of let-7 precursor (pre-let-7). But the enzyme responsible for the uridylation remained unknown. Here we identify a noncanonical poly (A) polymerase, TUTase4 (TUT4), as the uridylyl transferase for pre-let-7. Lin28 recruits TUT4 to pre-let-7 by recognizing a tetra-nucleotide sequence motif (GGAG) in the terminal loop. TUT4 in turn adds an oligouridine tail to the pre-let-7, which blocks Dicer processing. Other miRNAs with the same sequence motif (miR-107, -143, and -200c) are regulated through the same mechanism. Knockdown of TUT4 and Lin28 reduces the level of stem cell markers, suggesting that they are required for stem cell maintenance. This study uncovers the role of TUT4 and Lin28 as specific suppressors of miRNA biogenesis, which has implications for stem cell research and cancer biology.

Pluripotency factors Lin28 and Oct4 identify a sub-population of stem cell-like cells in ovarian cancer

“…Lin28 is known to block the production of a group of microRNAs, including let-7 microRNAs, which have been implicated previously in the regulation of cell proliferation and differentiation (Bussing et al, 2008;Heo et al, 2009;Trabucchi et al, 2009). The observed growth inhibition following Lin28 repression is likely the result, at least in part, of a restoration in let-7 microRNA expression, which has been linked to a translational blockade of target mRNAs involved in promoting cell cycle progression (Bussing et al, 2008).…”

“…The co-expression of Lin28 and Oct4 in a sub-population of high-grade ovarian cancer cells is particularly intriguing, given that both genes have critical roles in the maintenance of pluripotency, and hence the production of induced pluripotent stem cells (Yu, et al, 2007;Heo et al, 2009;Pei, 2009). Importantly, we have found that Lin28 specifically binds to Oct4 mRNA and stimulates its translation in both human ES cells and embryonic carcinoma PA-1 cells (Qiu et al, 2009).…”

“…It has been shown to block the processing of let-7 microRNAs (Heo et al, 2008;Newman et al, 2008;Rybak et al, 2008;Viswanathan et al, 2008) as well as a handful of other microRNAs (Heo et al, 2009;Trabucchi et al, 2009). In addition, Lin28 has been reported by us and others to bind to a specific subset of mRNAs, including those for IGF-2, Oct4, and several cell cycle-related factors, thereby modulating their translation (Polesskaya et al, 2007;Qiu et al, 2009;.…”

Identification of Loop Nucleotide Polymorphisms Affecting MicroRNA Processing and Function

“…Our findings strongly support the latter model, and suggest that further analyses are required to precisely dissect the role of the pri-miRNA loop in miRNA processing in vivo. Although several regulatory proteins have been demonstrated to bind to pri/pre-miRNA loops and affect the production of mature miRNA (Heo et al, 2008;2009;Michlewski and Caceres, 2010;Newman et al, 2008;Trabucchi et al, 2009;Viswanathan et al, 2008) and conserved loop sequences are required for the interaction between these regulatory factors (Heo et al, 2009;Newman et al, 2008), none of these proteins have been shown to be involved in the regulation of the processing accuracy of the mature miRNA 5′ seed region. Additionally, 5′ end polymorphisms have been detected in miRNA profiles of CD8 T cells (Wu et al, 2007), but there is little evidence to explain the heterogeneity of the 5′ end of mature miRNAs.…”

“…The crystal structure of mouse Lin-28 revealed that two folded domains of Lin-28 recognize two distinct regions of the miRNA terminal loop regions and are sufficient to inhibit let-7 biogenesis in vivo (Nam et al, 2011). TUT4, a non-canonical polyA polymerase, can be recruited by Lin-28 to precursor let-7 to block Dicer processing through premiRNA uridylation (Heo et al, 2009). Heteronuclear ribonucleoprotein A1 (hnRNP A1), a negative regulator of let-7a, binds the conserved terminal loop of pri-let-7a-1 and inhibits its processing by Drosha (Michlewski and Caceres, 2010).…”