Transcriptome-wide RNA Binding Analysis of C9orf72 Poly(PR) Dipeptides

Overview

Journal Life Sci Alliance

Specialties Biochemistry
Biology
Cell Biology
Molecular Biology

Date 2023 Jul 12

PMID 37438085

Authors

Rubika Balendra

Igor Ruiz de Los Mozos

Hana M Odeh

Idoia Glaria

Carmelo Milioto

Katherine M Wilson

Agnieszka M Ule

Martina Hallegger

Laura Masino

Stephen Martin

Rickie Patani

James Shorter

Jernej Ule

Adrian M Isaacs

Affiliations

Soon will be listed here.

Abstract

An intronic GGGGCC repeat expansion in is a common genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia. The repeats are transcribed in both sense and antisense directions to generate distinct dipeptide repeat proteins, of which poly(GA), poly(GR), and poly(PR) have been implicated in contributing to neurodegeneration. Poly(PR) binding to RNA may contribute to toxicity, but analysis of poly(PR)-RNA binding on a transcriptome-wide scale has not yet been carried out. We therefore performed crosslinking and immunoprecipitation (CLIP) analysis in human cells to identify the RNA binding sites of poly(PR). We found that poly(PR) binds to nearly 600 RNAs, with the sequence GAAGA enriched at the binding sites. In vitro experiments showed that poly(GAAGA) RNA binds poly(PR) with higher affinity than control RNA and induces the phase separation of poly(PR) into condensates. These data indicate that poly(PR) preferentially binds to poly(GAAGA)-containing RNAs, which may have physiological consequences.

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References

Moens T, Niccoli T, Wilson K, Atilano M, Birsa N, Gittings L . C9orf72 arginine-rich dipeptide proteins interact with ribosomal proteins in vivo to induce a toxic translational arrest that is rescued by eIF1A. Acta Neuropathol. 2019; 137(3):487-500. PMC: 6514073. DOI: 10.1007/s00401-018-1946-4. View

Kramer N, Haney M, Morgens D, Jovicic A, Couthouis J, Li A . CRISPR-Cas9 screens in human cells and primary neurons identify modifiers of C9ORF72 dipeptide-repeat-protein toxicity. Nat Genet. 2018; 50(4):603-612. PMC: 5893388. DOI: 10.1038/s41588-018-0070-7. View

Moens T, Partridge L, Isaacs A . Genetic models of C9orf72: what is toxic?. Curr Opin Genet Dev. 2017; 44:92-101. DOI: 10.1016/j.gde.2017.01.006. View

Kwon I, Xiang S, Kato M, Wu L, Theodoropoulos P, Wang T . Poly-dipeptides encoded by the C9orf72 repeats bind nucleoli, impede RNA biogenesis, and kill cells. Science. 2014; 345(6201):1139-45. PMC: 4459787. DOI: 10.1126/science.1254917. View

DeJesus-Hernandez M, Mackenzie I, Boeve B, Boxer A, Baker M, Rutherford N . Expanded GGGGCC hexanucleotide repeat in noncoding region of C9ORF72 causes chromosome 9p-linked FTD and ALS. Neuron. 2011; 72(2):245-56. PMC: 3202986. DOI: 10.1016/j.neuron.2011.09.011. View

Zu T, Liu Y, Banez-Coronel M, Reid T, Pletnikova O, Lewis J . RAN proteins and RNA foci from antisense transcripts in C9ORF72 ALS and frontotemporal dementia. Proc Natl Acad Sci U S A. 2013; 110(51):E4968-77. PMC: 3870665. DOI: 10.1073/pnas.1315438110. View

Mizielinska S, Gronke S, Niccoli T, Ridler C, Clayton E, Devoy A . C9orf72 repeat expansions cause neurodegeneration in Drosophila through arginine-rich proteins. Science. 2014; 345(6201):1192-1194. PMC: 4944841. DOI: 10.1126/science.1256800. View

Lee K, Zhang P, Kim H, Mitrea D, Sarkar M, Freibaum B . C9orf72 Dipeptide Repeats Impair the Assembly, Dynamics, and Function of Membrane-Less Organelles. Cell. 2016; 167(3):774-788.e17. PMC: 5079111. DOI: 10.1016/j.cell.2016.10.002. View

Nussbacher J, Tabet R, Yeo G, Lagier-Tourenne C . Disruption of RNA Metabolism in Neurological Diseases and Emerging Therapeutic Interventions. Neuron. 2019; 102(2):294-320. PMC: 6545120. DOI: 10.1016/j.neuron.2019.03.014. View

10.

Lin Y, Mori E, Kato M, Xiang S, Wu L, Kwon I . Toxic PR Poly-Dipeptides Encoded by the C9orf72 Repeat Expansion Target LC Domain Polymers. Cell. 2016; 167(3):789-802.e12. PMC: 5076566. DOI: 10.1016/j.cell.2016.10.003. View

11.

Huppertz I, Attig J, DAmbrogio A, Easton L, Sibley C, Sugimoto Y . iCLIP: protein-RNA interactions at nucleotide resolution. Methods. 2013; 65(3):274-87. PMC: 3988997. DOI: 10.1016/j.ymeth.2013.10.011. View

12.

Dominguez D, Freese P, Alexis M, Su A, Hochman M, Palden T . Sequence, Structure, and Context Preferences of Human RNA Binding Proteins. Mol Cell. 2018; 70(5):854-867.e9. PMC: 6062212. DOI: 10.1016/j.molcel.2018.05.001. View

13.

Yin S, Lopez-Gonzalez R, Kunz R, Gangopadhyay J, Borufka C, Gygi S . Evidence that C9ORF72 Dipeptide Repeat Proteins Associate with U2 snRNP to Cause Mis-splicing in ALS/FTD Patients. Cell Rep. 2017; 19(11):2244-2256. PMC: 5653973. DOI: 10.1016/j.celrep.2017.05.056. View

14.

Patel A, Lee H, Jawerth L, Maharana S, Jahnel M, Hein M . A Liquid-to-Solid Phase Transition of the ALS Protein FUS Accelerated by Disease Mutation. Cell. 2015; 162(5):1066-77. DOI: 10.1016/j.cell.2015.07.047. View

15.

Freibaum B, Lu Y, Lopez-Gonzalez R, Kim N, Almeida S, Lee K . GGGGCC repeat expansion in C9orf72 compromises nucleocytoplasmic transport. Nature. 2015; 525(7567):129-33. PMC: 4631399. DOI: 10.1038/nature14974. View

16.

Renton A, Majounie E, Waite A, Simon-Sanchez J, Rollinson S, Gibbs J . A hexanucleotide repeat expansion in C9ORF72 is the cause of chromosome 9p21-linked ALS-FTD. Neuron. 2011; 72(2):257-68. PMC: 3200438. DOI: 10.1016/j.neuron.2011.09.010. View

17.

Hartmann H, Hornburg D, Czuppa M, Bader J, Michaelsen M, Farny D . Proteomics and neuropathology identify ribosomes as poly-GR/PR interactors driving toxicity. Life Sci Alliance. 2018; 1(2):e201800070. PMC: 6238541. DOI: 10.26508/lsa.201800070. View

18.

Zhang Y, Gendron T, Ebbert M, ORaw A, Yue M, Jansen-West K . Poly(GR) impairs protein translation and stress granule dynamics in C9orf72-associated frontotemporal dementia and amyotrophic lateral sclerosis. Nat Med. 2018; 24(8):1136-1142. PMC: 6520050. DOI: 10.1038/s41591-018-0071-1. View

19.

Lopez-Gonzalez R, Lu Y, Gendron T, Karydas A, Tran H, Yang D . Poly(GR) in C9ORF72-Related ALS/FTD Compromises Mitochondrial Function and Increases Oxidative Stress and DNA Damage in iPSC-Derived Motor Neurons. Neuron. 2016; 92(2):383-391. PMC: 5111366. DOI: 10.1016/j.neuron.2016.09.015. View

20.

Haeusler A, Donnelly C, Periz G, Simko E, Shaw P, Kim M . C9orf72 nucleotide repeat structures initiate molecular cascades of disease. Nature. 2014; 507(7491):195-200. PMC: 4046618. DOI: 10.1038/nature13124. View