Fig. 6

Mitochondrial respiration, ER-mitochondrial contacts, and intracellular calcium handling in HEK293T cells expressing mutant TDP-43. a Western blot of cell homogenates from HEK293T cells transiently transfected with constructs expressing recombinant TDP-43-myc probed with myc and GAPDH as a loading control. b Quantification of TDP-43 band intensities relative to the intensity of GAPDH (average of two western blots). c OCR baseline measurements of HEK293T cells transfected with TDP-43-myc constructs. All lines were tested in 12 technical over three biological replicates. d Representative micrographs of HeLa cells co-transfected with the fluorescent reporters ddGFP and mitoDSRed. e Mean co-localization of mitoDSRed and ddGFP in HeLa cells expressing WT or mutant TDP-43. n = 30–40 cells over three independent transfections. f Representative micrograph of HeLa cells transfected with the fluorescent reporter GCamp6 targeted to mitochondria (mtGCamp6). g Average mtGCamp6 fluorescence curves indicating mitochondrial calcium dynamics in HeLa cells expressing WT or mutant TDP-43 in response to mobilization of ER calcium stores induced by perfusion with ATP in calcium free buffer, followed by perfusion with 2 mM calcium buffer to induce SOCE. n = 12–15 cells over two independent transfections