Argonaute Utilization for miRNA Silencing Is Determined by Phosphorylation-Dependent Recruitment of LIM-Domain-Containing Proteins

Katherine S Bridge; Kunal M Shah; Yigen Li; Daniel E Foxler; Sybil C K Wong; Duncan C Miller; Kathryn M Davidson; John G Foster; Ruth Rose; Michael R Hodgkinson; Paulo S Ribeiro; A Aziz Aboobaker; Kenta Yashiro; Xiaozhong Wang; Paul R Graves; Michael J Plevin; Dimitris Lagos; Tyson V Sharp

doi:10.1016/j.celrep.2017.06.027

Argonaute Utilization for miRNA Silencing Is Determined by Phosphorylation-Dependent Recruitment of LIM-Domain-Containing Proteins

Cell Rep. 2017 Jul 5;20(1):173-187. doi: 10.1016/j.celrep.2017.06.027.

Authors

Katherine S Bridge¹, Kunal M Shah¹, Yigen Li¹, Daniel E Foxler¹, Sybil C K Wong¹, Duncan C Miller¹, Kathryn M Davidson¹, John G Foster¹, Ruth Rose², Michael R Hodgkinson³, Paulo S Ribeiro⁴, A Aziz Aboobaker⁵, Kenta Yashiro⁶, Xiaozhong Wang⁷, Paul R Graves⁸, Michael J Plevin³, Dimitris Lagos⁹, Tyson V Sharp¹⁰

Affiliations

¹ Centre for Molecular Oncology, Barts Cancer Institute, Queen Mary University of London, John Vane Science Centre, Charterhouse Square, London EC1M 6BQ, UK.
² School of Biological and Chemical Sciences, Queen Mary University of London, Fogg Building, Mile End Road, London E1 4NS, UK.
³ Department of Biology, University of York, Heslington, York YO10 5DD, UK.
⁴ Centre for Tumour Biology, Barts Cancer Institute, Queen Mary University of London, John Vane Science Centre, Charterhouse Square, London EC1M 6BQ, UK.
⁵ Department of Zoology, University of Oxford, The Tinbergen Building, South Parks Road, Oxford OX1 3PS, UK.
⁶ Cardiac Regeneration and Therapeutics, Osaka University Graduate School of Medicine, 2-2 Yamada-oka, Suita, Osaka 565-0871, Japan.
⁷ Department of Biochemistry, Molecular Biology and Cell Biology, Northwestern University, Evanston, IL 60208, USA.
⁸ Department of Radiation Oncology, New York-Presbyterian Brooklyn Methodist Hospital, 506 6th Street, Brooklyn, NY 11215, USA.
⁹ Centre for Immunology and Infection, Hull York Medical School and Department of Biology, University of York, Heslington, York YO10 5DD, UK.
¹⁰ Centre for Molecular Oncology, Barts Cancer Institute, Queen Mary University of London, John Vane Science Centre, Charterhouse Square, London EC1M 6BQ, UK. Electronic address: t.sharp@qmul.ac.uk.

Abstract

As core components of the microRNA-induced silencing complex (miRISC), Argonaute (AGO) proteins interact with TNRC6 proteins, recruiting other effectors of translational repression/mRNA destabilization. Here, we show that LIMD1 coordinates the assembly of an AGO-TNRC6 containing miRISC complex by binding both proteins simultaneously at distinct interfaces. Phosphorylation of AGO2 at Ser 387 by Akt3 induces LIMD1 binding, which in turn enables AGO2 to interact with TNRC6A and downstream effector DDX6. Conservation of this serine in AGO1 and 4 indicates this mechanism may be a fundamental requirement for AGO function and miRISC assembly. Upon CRISPR-Cas9-mediated knockout of LIMD1, AGO2 miRNA-silencing function is lost and miRNA silencing becomes dependent on a complex formed by AGO3 and the LIMD1 family member WTIP. The switch to AGO3 utilization occurs due to the presence of a glutamic acid residue (E390) on the interaction interface, which allows AGO3 to bind to LIMD1, AJUBA, and WTIP irrespective of Akt signaling.

Keywords: AGO2; AKT; Argonaute; DDX6; GW182; LIMD1; TNRC6A; WTIP; miRISC; microRNA.

MeSH terms

Argonaute Proteins / genetics
Argonaute Proteins / metabolism*
Autoantigens / metabolism
DEAD-box RNA Helicases / metabolism
Gene Silencing*
HEK293 Cells
HeLa Cells
Humans
Intracellular Signaling Peptides and Proteins / chemistry
Intracellular Signaling Peptides and Proteins / genetics
Intracellular Signaling Peptides and Proteins / metabolism*
LIM Domain Proteins / chemistry
LIM Domain Proteins / genetics
LIM Domain Proteins / metabolism*
MicroRNAs / genetics*
MicroRNAs / metabolism
Phosphorylation
Protein Binding
Protein Processing, Post-Translational
Proto-Oncogene Proteins / metabolism
RNA-Binding Proteins / metabolism

Substances

AGO2 protein, human
Argonaute Proteins
Autoantigens
Intracellular Signaling Peptides and Proteins
LIM Domain Proteins
LIMD1 protein, human
MicroRNAs
Proto-Oncogene Proteins
RNA-Binding Proteins
TNRC6A protein, human
DDX6 protein, human
DEAD-box RNA Helicases

Abstract

MeSH terms

Substances

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