C9orf72 FTLD/ALS-associated Gly-Ala dipeptide repeat proteins cause neuronal toxicity and Unc119 sequestration

Acta Neuropathol. 2014 Oct;128(4):485-503. doi: 10.1007/s00401-014-1329-4. Epub 2014 Aug 14.


Hexanucleotide repeat expansion in C9orf72 is the most common pathogenic mutation in patients with amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). Despite the lack of an ATG start codon, the repeat expansion is translated in all reading frames into dipeptide repeat (DPR) proteins, which form insoluble, ubiquitinated, p62-positive aggregates that are most abundant in the cerebral cortex and cerebellum. To specifically analyze DPR toxicity and aggregation, we expressed DPR proteins from synthetic genes containing a start codon but lacking extensive GGGGCC repeats. Poly-Gly-Ala (GA) formed p62-positive cytoplasmic aggregates, inhibited dendritic arborization and induced apoptosis in primary neurons. Quantitative mass spectrometry analysis to identify poly-GA co-aggregating proteins revealed a significant enrichment of proteins of the ubiquitin-proteasome system. Among the other interacting proteins, we identified the transport factor Unc119, which has been previously linked to neuromuscular and axonal function, as a poly-GA co-aggregating protein. Strikingly, the levels of soluble Unc119 are strongly reduced upon poly-GA expression in neurons, suggesting a loss of function mechanism. Similar to poly-GA expression, Unc119 knockdown inhibits dendritic branching and causes neurotoxicity. Unc119 overexpression partially rescues poly-GA toxicity suggesting that poly-GA expression causes Unc119 loss of function. In C9orf72 patients, Unc119 is detectable in 9.5 % of GA inclusions in the frontal cortex, but only in 1.6 % of GA inclusions in the cerebellum, an area largely spared of neurodegeneration. A fraction of neurons with Unc119 inclusions shows loss of cytosolic staining. Poly-GA-induced Unc119 loss of function may thereby contribute to selective vulnerability of neurons with DPR protein inclusions in the pathogenesis of C9orf72 FTLD/ALS.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptor Proteins, Signal Transducing / metabolism*
  • Amyotrophic Lateral Sclerosis / genetics
  • Amyotrophic Lateral Sclerosis / metabolism*
  • Amyotrophic Lateral Sclerosis / pathology
  • Animals
  • Caspases / metabolism
  • Cells, Cultured
  • Cerebral Cortex / cytology
  • DNA Repeat Expansion / genetics
  • DNA Repeat Expansion / physiology
  • DNA-Binding Proteins / metabolism
  • Dipeptides / genetics
  • Dipeptides / metabolism*
  • Dipeptides / pharmacology
  • Frontotemporal Lobar Degeneration / genetics
  • Frontotemporal Lobar Degeneration / metabolism*
  • Frontotemporal Lobar Degeneration / pathology
  • Gene Expression Regulation / drug effects
  • Gene Expression Regulation / genetics
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • HEK293 Cells
  • Humans
  • Microtubule-Associated Proteins / metabolism
  • RNA-Binding Proteins / chemistry*
  • RNA-Binding Proteins / genetics
  • RNA-Binding Proteins / metabolism*
  • Rats
  • Transfection
  • Ubiquitin / metabolism


  • Adaptor Proteins, Signal Transducing
  • DNA-Binding Proteins
  • Dipeptides
  • FAM120A protein, human
  • MAP2 protein, human
  • Microtubule-Associated Proteins
  • P62 protein, human
  • RNA-Binding Proteins
  • UNC119 protein, human
  • Ubiquitin
  • Green Fluorescent Proteins
  • N-glycylalanine
  • Caspases