C9orf72 and triplet repeat disorder RNAs: G-quadruplex formation, binding to PRC2 and implications for disease mechanisms

RNA. 2019 Aug;25(8):935-947. doi: 10.1261/rna.071191.119. Epub 2019 May 2.


Some neurological disorders, including amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), fragile X syndrome, Huntington's disease, myotonic dystrophy, and various ataxias, can be caused by expansions of short nucleic acid sequence repeats in specific genes. A possible disease mechanism involves the transcribed repeat RNA binding an RNA-binding protein (RBP), resulting in its sequestration and thus dysfunction. Polycomb repressive complex 2 (PRC2), the histone methyltransferase that deposits the H3K27me3 mark of epigenetically silenced chromatin, binds G-rich RNAs and has especially high affinity for G-quadruplex (G-Q) structures. Here, we find that PRC2 target genes are derepressed and the RNA binding subunit EZH2 largely insoluble in postmortem brain samples from ALS/FTD patients with C9ORF72 (C9) repeat expansions, leading to the hypothesis that the (G4C2)n repeat RNA might be sequestering PRC2. Contrary to this expectation, we found that C9 repeat RNAs (n = 6 or 10) bind weakly to purified PRC2, and studies with the G-Q specific BG4 antibody and circular dichroism studies both indicated that these C9 RNAs have little propensity to form G-Qs in vitro. Several GC-rich triplet-repeat expansion RNAs also have low affinity for PRC2 and do not appreciably form G-Qs in vitro. The results are consistent with these sequences forming hairpin structures that outcompete G-Q folding when the repeat length is sufficiently large. We suggest that binding of PRC2 to these GC-rich RNAs is fundamentally weak but may be modulated in vivo by protein factors that affect secondary structure, such as helicases and other RBPs.

Keywords: ALS; G-quadruplex; RNA–protein interactions; epigenetics; neurological disorders.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amyotrophic Lateral Sclerosis / genetics*
  • Amyotrophic Lateral Sclerosis / metabolism
  • Autopsy
  • C9orf72 Protein / chemistry*
  • C9orf72 Protein / genetics*
  • Circular Dichroism
  • Enhancer of Zeste Homolog 2 Protein / chemistry
  • Enhancer of Zeste Homolog 2 Protein / metabolism
  • Epigenesis, Genetic
  • Frontotemporal Dementia / genetics*
  • Frontotemporal Dementia / metabolism
  • G-Quadruplexes
  • Humans
  • Polycomb Repressive Complex 2 / chemistry
  • Polycomb Repressive Complex 2 / metabolism*
  • Solubility
  • Trinucleotide Repeats*


  • C9orf72 Protein
  • EZH2 protein, human
  • Enhancer of Zeste Homolog 2 Protein
  • Polycomb Repressive Complex 2

Supplementary concepts

  • Frontotemporal Dementia With Motor Neuron Disease