Figure 8.

T209K-Gars has altered sub-cellular localization. (A) Expression of wild-type and mutant GARS-EGFP in mouse motor neurons. The T130K mutation was introduced into the human-coding sequence in frame with a C-terminal EGFP tag, and transfected into MN-1 cells. Cells expressing wild-type GARS-EGFP demonstrated localization of the protein to cytoplasmic puncta (arrowhead). (B) T130K GARS-EGFP appeared diffused throughout the cytoplasm and did not form punctate structures. (C and D) Confocal analysis in agarose sections width 50 μm of 96 hpf wild-type and gars^s266/s266 larvae stained with filamentous actin (red), anti-Gars (green) and the ac-a-tubulin (blue), a pan-axonal marker (wt n = 12, gars^s266/s266 n = 11). (C) Endogenous Gars is localized in cytoplasmic puncta of the neurons in the midbrain of wild-type larvae at 96 hpf. (D) In contrast, in mutant larvae Gars showed fainter, diffused staining. Scale bars: 5 μm.