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. 2016 Apr 21;35(12):1276–1297. doi: 10.15252/embj.201593350

Figure EV5. Reduced expression of C9ORF72 synergizes Ataxin‐2 toxicity.

Figure EV5

  • A
    Representative images of GT1‐7 neuronal cells co‐transfected with HA‐tagged ATXN2 with control (Q22x) or intermediate (Q30x) polyQ size and either control siRNA or siRNA targeting endogenous C9orf72 mRNA.
  • B, C
    Immunoblot analysis of endogenous Gapdh or transfected HA‐tagged Ataxin‐2 with (B) control (Q22x) length or (C) intermediate (Q30x) length of polyQ in GT1‐7 cells.
  • D
    Representative images of GT1‐7 neuronal cells co‐transfected with either GFP‐tagged mutant SOD1, FUS, HTT, or Ataxin‐3 and either control siRNA or siRNA targeting endogenous C9orf72 mRNA.
  • E
    RT–qPCR quantification of endogenous C9orf72 expression relative to Gapdh mRNA in mismatched or C9orf72 antisense morpholino oligonucleotide (AMO)‐injected zebrafish.
  • F
    RT–qPCR quantification of exogenous HA‐tagged ATXN2 with normal (Q22x) or intermediate (Q30x) length of polyQ relative to endogenous Gapdh mRNA in zebrafish injected with control or HA‐tagged ATXN2 constructs and with mismatched AMOs or AMOs against C9orf72.
  • G
    Tracing of the swimming trajectories of 48 h post‐fertilization zebrafish larvae following light touch.
  • H–J
    Quantification of the touch‐evoked swimming distance (H), average velocity (I), and maximum velocity attained (J) shows no impairment upon the sole injection of HA‐tagged ATXN2 with control (Q22x) or intermediate length of polyQ (Q30x). Similarly, injection of mismatched antisense morpholino oligonucleotides (AMOs) against C9orf72 alone or with HA‐tagged ATXN2 Q22x or Q30x shows no functional alterations.

Data information: Scale bars, 10 μm. Nuclei were counterstained with DAPI. Error bars indicate SEM, n = 3.