Fig. 1

Aβ induces mitochondria fragmentation by promoting Drp1 O-GlcNAcylation in neuronal cells. a Primary cultured neuron were exposed to control (Con), Aβ (10 µM), or PugNAc (100 µM). Mitochondria were stained with mitotracker-Red. The mean mitochondrial length was determined from images of multiple cells obtained by microscopy. The bar graph indicates means ± SEM. *p < 0.01. The scale bar indicates 10 µm. b Drp1 O-GlcNAcylation in response to either Aβ (10 µM) or PugNAc (100 µM) in SH-SY5Y. Cell lysates were immunoprecipitated (IP) with anti-WGA-agarose antibody and immunoblotted with anti-Drp1 antibody. c Drp1 O-GlcNAcylation by Aβ treatment in primary neuron. d Enhanced Drp1 O-GlcNAcylation in brain tissue of 5 × FAD. Mice brain lysates from wild-type and 5 × FAD were assessed by IP using anti-WGA-agarose antibody and confirmed with anti-Drp1 and anti-Nicastrin antibodies. e Depletion of OGT reduced O-GlcNAcylation of Drp1. SH-SY5Y cells transfected with scrambled siRNA (Sc) or siRNA for OGT were treated with Aβ (10 µM) and the cell lysates were assessed by IP with anti-WGA-agarose antibody and immunoblotted with anti-Drp1. f Interaction between Drp1 and OGT. SH-SY5Y expressing OGT-flag and GFP-Drp1 were treated with Aβ (10 µM) and then assessed with IP (anti-GFP antibody) and immunoblotted with anti-Flag and anti-GFP antibodies. g O-GlcNAcylation of mutant Drp1. SH-SY5Y cells expressing empty vector (Vec), GFP-Drp1 (Drp1), or GFP-Drp1 T585A/586A mutant (Drp1 TA) were treated with Aβ (10 µM). The cell lysates were assessed by IP with anti-WGA-agarose antibody and immunoblotted with anti-GFP-antibody. h Mitochondrial fragmentation with Drp1 mutant. SH-SY5Y cells expressing empty vector (Vec), GFP-Drp1 (Drp1), or GFP-Drp1 T585A/586A mutant (Drp1 TA) were treated with Aβ (10 µM). Cells with fragmented mitochondria were counted under a microscopy. The bar graph indicates means ± SEM. *p < 0.01