microRNA-Mediated Messenger RNA Deadenylation Contributes to Translational Repression in Mammalian Cells

Abstract

Animal microRNAs (miRNAs) typically regulate gene expression by binding to partially complementary target sites in the 3′ untranslated region (UTR) of messenger RNA (mRNA) reducing its translation and stability. They also commonly induce shortening of the mRNA 3′ poly(A) tail, which contributes to their mRNA decay promoting function. The relationship between miRNA-mediated deadenylation and translational repression has been less clear. Using transfection of reporter constructs carrying three imperfectly matching let-7 target sites in the 3′ UTR into mammalian cells we observe rapid target mRNA deadenylation that precedes measureable translational repression by endogenous let-7 miRNA. Depleting cells of the argonaute co-factors RCK or TNRC6A can impair let-7-mediated repression despite ongoing mRNA deadenylation, indicating that deadenylation alone is not sufficient to effect full repression. Nevertheless, the magnitude of translational repression by let-7 is diminished when the target reporter lacks a poly(A) tail. Employing an antisense strategy to block deadenylation of target mRNA with poly(A) tail also partially impairs translational repression. On the one hand, these experiments confirm that tail removal by deadenylation is not strictly required for translational repression. On the other hand they show directly that deadenylation can augment miRNA-mediated translational repression in mammalian cells beyond stimulating mRNA decay. Taken together with published work, these results suggest a dual role of deadenylation in miRNA function: it contributes to translational repression as well as mRNA decay and is thus critically involved in establishing the quantitatively appropriate physiological response to miRNAs.

FXR1a-associated microRNP: A driver of specialized non-canonical translation in quiescent conditions

“…1,11,76,77 Only unadenylated or poly(A) shortened (shorter than a PABP site) reporter mRNAs were translationally upregulated in the presence of the microRNA in G0 THP1 cells and in immature oocytes. 31 Measurement of poly(A) tail length of the validated endogenous targets of activation, using poly(A) tail assay, showed shortened poly(A) tails of these targets.…”

Post-Transcriptional Regulation of DNA Damage-Responsive Gene Expression

“…5). Sequence-specific binding of miRNAs to target mRNAs can direct the transcript for deadenylation-dependent decay and/or translational inhibition (5,137).…”

The Role of microRNAs in Gliomas and Their Potential Applications for Diagnosis and Treatment

“…A second model has proposed that Ago recruits eIF6 and, thus, prevents the association of the 60S ribosomal subunit with the 40S preinitiation complex 28,29 . The third model suggests that miRNAs destabilize target mRNAs by de-adenylation and subsequent decapping [30][31][32][33][34] . It was recently observed that AGO-2, mature miRNAs and translationally repressed mRNAs can accumulate in cytoplasmic processing bodies (P-bodies) 35 .…”