Drosophila lines with mutant and wild type human TDP-43 replacing the endogenous gene reveals phosphorylation and ubiquitination in mutant lines in the absence of viability or lifespan defects

PLoS One. 2017 Jul 7;12(7):e0180828. doi: 10.1371/journal.pone.0180828. eCollection 2017.

Abstract

Mutations in TDP-43 are associated with proteinaceous inclusions in neurons and are believed to be causative in neurodegenerative diseases such as frontotemporal dementia or amyotrophic lateral sclerosis. Here we describe a Drosophila system where we have engineered the genome to replace the endogenous TDP-43 orthologue with wild type or mutant human TDP-43(hTDP-43). In contrast to other models, these flies express both mutant and wild type hTDP-43 at similar levels to those of the endogenous gene and importantly, no age-related TDP-43 accumulation observed among all the transgenic fly lines. Immunoprecipitation of TDP-43 showed that flies with hTDP-43 mutations had increased levels of ubiquitination and phosphorylation of the hTDP-43 protein. Furthermore, histologically, flies expressing hTDP-43 M337V showed global, robust neuronal staining for phospho-TDP. All three lines: wild type hTDP-43, -G294A and -M337V were homozygous viable, with no defects in development, life span or behaviors observed. The primary behavioral defect was that flies expressing either hTDP-43 G294A or M337V showed a faster decline with age in negative geotaxis. Together, these observations implied that neurons could handle these TDP-43 mutations by phosphorylation- and ubiquitin-dependent proteasome systems, even in a background without the wild type TDP-43. Our findings suggest that these two specific TDP-43 mutations are not inherently toxic, but may require additional environmental or genetic factors to affect longevity or survival.

MeSH terms

  • Aging / metabolism*
  • Animals
  • Animals, Genetically Modified
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism*
  • Drosophila melanogaster / genetics*
  • Drosophila melanogaster / growth & development
  • Drosophila melanogaster / metabolism
  • Female
  • Humans
  • Longevity*
  • Male
  • Mutation
  • Neurons / cytology
  • Neurons / metabolism*
  • Phosphorylation
  • Protein Processing, Post-Translational*
  • Signal Transduction
  • Taxis Response / physiology
  • Transgenes
  • Ubiquitination

Substances

  • DNA-Binding Proteins
  • TARDBP protein, human