Let-7 microRNA-mediated mRNA deadenylation and translational repression in a mammalian cell-free system

• Published in: Genes & development Vol. 21; no. 15; pp. 1857 - 1862
• Main Authors: Wakiyama, Motoaki, Takimoto, Koji, Ohara, Osamu, Yokoyama, Shigeyuki
• Format: Journal Article
• Language: English
• Published: United States Cold Spring Harbor Laboratory Press 01.08.2007
• Subjects: Argonaute Proteins; Autoantigens - genetics; Base Sequence; Cell Line; Cell-Free System; Eukaryotic Initiation Factor-2 - genetics; Gene Silencing; Genes, Reporter; Humans; In Vitro Techniques; Luciferases, Firefly - genetics; MicroRNAs - genetics; MicroRNAs - metabolism; Polyadenylation; Protein Biosynthesis; Research Communication; RNA Caps - chemistry; RNA Caps - genetics; RNA Caps - metabolism; RNA, Messenger - chemistry; RNA, Messenger - genetics; RNA, Messenger - metabolism; RNA, Small Interfering - genetics; RNA-Binding Proteins
• ISSN: 0890-9369; 1549-5477
• DOI: 10.1101/gad.1566707

MicroRNAs (miRNAs) are incorporated into miRNP complexes and regulate protein expression post-transcriptionally through binding to 3'-untranslated regions of target mRNAs. Here we describe a recapitulation of let-7 miRNA-mediated translational repression in a cell-free system, which was established with extracts prepared from HEK293F cells overexpressing miRNA pathway components. In this system, both the cap and poly(A) tail are required for the translational repression, and let-7 directs the deadenylation of target mRNAs. Our work suggests that let-7 miRNPs containing Argonaute and GW182 impair the synergistic enhancement of translation by the 5'-cap and 3'-poly(A) tail, resulting in translational repression.