Overexpression of Toxic Poly(Glycine-Alanine) Aggregates in Primary Neuronal Cultures Induces Time-Dependent Autophagic and Synaptic Alterations but Subtle Activity Impairments

Cells. 2024 Aug 3;13(15):1300. doi: 10.3390/cells13151300.

Abstract

The pathogenic expansion of the intronic GGGGCC hexanucleotide located in the non-coding region of the C9orf72 gene represents the most frequent genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). This mutation leads to the accumulation of toxic RNA foci and dipeptide repeats (DPRs), as well as reduced levels of the C9orf72 protein. Thus, both gain and loss of function are coexisting pathogenic aspects linked to C9orf72-ALS/FTD. Synaptic alterations have been largely described in C9orf72 models, but it is still not clear which aspect of the pathology mostly contributes to these impairments. To address this question, we investigated the dynamic changes occurring over time at the synapse upon accumulation of poly(GA), the most abundant DPR. Overexpression of this toxic form induced a drastic loss of synaptic proteins in primary neuron cultures, anticipating autophagic defects. Surprisingly, the dramatic impairment characterizing the synaptic proteome was not fully matched by changes in network properties. In fact, high-density multi-electrode array analysis highlighted only minor reductions in the spike number and firing rate of poly(GA) neurons. Our data show that the toxic gain of function linked to C9orf72 affects the synaptic proteome but exerts only minor effects on the network activity.

Keywords: ALS; C9orf72; MEA; autophagy; poly(GA); synapse.

MeSH terms

  • Animals
  • Autophagy*
  • C9orf72 Protein* / genetics
  • C9orf72 Protein* / metabolism
  • Cells, Cultured
  • Humans
  • Neurons* / metabolism
  • Peptides / metabolism
  • Protein Aggregates
  • Synapses* / metabolism

Substances

  • C9orf72 Protein
  • Peptides
  • Protein Aggregates

Grants and funding

This research was funded by the Deutsche Forschungsgemeinschaft (DFG; German Research Foundation) through the SFB 1506-Project 01 to AC, who receives further support from the Frick Foundation for ALS research, the Else Kröner-Fresenius Stiftung (project Nr. 2019_A111; L.SBN.0162), the German Society for Muscle Disease (DGM; project Ca2/1), the Karin Christiane Conradi Foundation, the Deutsche Forschungsgemeinschaft (individual project CA 2915/4-1), and the Medical Scientist Program of the Ulm Medical Faculty.