Let‐7a‐regulated translational readthrough of mammalian AGO 1 generates a micro RNA pathway inhibitor

Abstract

Translational readthrough generates proteins with extended C‐termini, which often possess distinct properties. Here, we have used various reporter assays to demonstrate translational readthrough of AGO1 mRNA. Analysis of ribosome profiling data and mass spectrometry data provided additional evidence for translational readthrough of AGO1. The endogenous readthrough product, Ago1x, could be detected by a specific antibody both in vitro and in vivo. This readthrough process is directed by a cis sequence downstream of the canonical AGO1 stop codon, which is sufficient to drive readthrough even in a heterologous context. This cis sequence has a let‐7a miRNA‐binding site, and readthrough is promoted by let‐7a miRNA. Interestingly, Ago1x can load miRNAs on target mRNAs without causing post‐transcriptional gene silencing, due to its inability to interact with GW182. Because of these properties, Ago1x can serve as a competitive inhibitor of miRNA pathway. In support of this, we observed increased global translation in cells overexpressing Ago1x. Overall, our results reveal a negative feedback loop in the miRNA pathway mediated by the translational readthrough product of AGO1.

Stop codon readthrough alters the activity of a POU/Oct transcription factor during Drosophila development

“…Alternative decoding of factors that regulate gene expression could consequentially result in broad changes in downstream processes. For example, SCR of the mammalian gene for Argonaute1 (Ago1) produces the Ago1x isoform, which acts as a competitive inhibitor of the miRNA pathway, leading to increased global translation as a result of SCR [ 12 ]. In the present work, a deletion that abolished the C-terminal extension had strong effects on the transcriptional profile of dfr 14 mutant larvae, with pronounced effects on genes involved in gene expression, neural proliferation, sensory organ development, and immune system processes.…”

“…For example, heterogeneous nuclear ribonucleoprotein (hnRNP) A2/B1 was shown to bind to a cis -acting element in VEGF-A 3′ untranslated region (UTR) and promote SCR [ 11 ]. Translational readthrough of the mammalian AGO1 gene, encoding the Argonaute 1 (Ago1) protein, was recently found to be positively regulated by the let-7a miRNA upon binding 3′ of the canonical stop codon [ 12 ].…”

De novocoding variants in theAGO1gene cause a neurodevelopmental disorder with intellectual disability

“…Recently, the AGO1x protein isoform was described, product of the translational stop read-through of AGO 1 transcripts induced by the expression of the Let7a miRNA. Ectopic expression of AGO1x acts as a negative competitor of the miRNA pathway 36 . A hypothesis would be that mutated AGO1 proteins may act as the AGO1x isoform through competitive inhibition of wild type AGO proteins.…”

Argonaute Proteins: From Structure to Function in Development and Pathological Cell Fate Determination

“…Finally, AGO1 has also been detected as a candidate for programed translational readthrough, a process generating longer isoforms by continuing translation beyond the stop codon (Eswarappa et al, 2014). Two recent studies demonstrated the presence of this translational readthrough product of AGO1 in cells, termed AGO1x (Ghosh et al, 2019;Singh et al, 2019). AGO1x is a protein isoform, which is 33 amino acids longer than AGO1 (Figure 1A).…”

“…Since AGO1x is incapable of interacting with GW182, it cannot induce translational repression. It is therefore thought that AGO1x competes with the canonical miRNA pathway and thereby leads to reduced posttranscriptional repression of target mRNAs (Singh et al, 2019). Second, in breast cancer cells, AGO1x has been shown to prevent the accumulation of dsRNAs and thereby suppresses the interferon response in these cells, a function independent of the canonical miRNA pathway (Ghosh et al, 2019).…”