Figure 2. PINK1-SPARK characterizes PINK1 activity in multiple cell types.

A. Left: representative images of HeLa, undifferentiated SH-SY5Y (USH), and neuronally differentiated SH-SY5Y (DSH) cells expressing PINK1-SPARK when treated with DMSO or 10 μM CCCP. Right: representative images of various cell types expressing the phosphonull mutant PINK1-SPARK (S/A) following addition of DMSO or CCCP.

B. Number of puncta per cell in either WT HeLa or PINK1 KO HeLa cell following treatment with DMSO (WT: n = 141 cells from 3 experiments; PINK1 KO: n = 236 cells from 3 experiments) or 10 μM CCCP (WT: n = 159 cells from 4 experiments; PINK1 KO: n = 230 cells from 3 experiments; ****p < 0.0001; ns = 0.3962, unpaired t-test, two-tailed).

C. Percentage of WT or PINK1 KO HeLa cells with puncta following addition of DMSO or 10 μM CCCP (**p = 0.0014; unpaired t-test, two-tailed).

D. Puncta area in HeLa cells when treated with 10 μM CCCP (n = 463 puncta from 3 experiments).

E. Number of puncta per USH cell following treatment with DMSO (n = 94 cells from 3 experiments) or 10 μM CCCP (n = 27 cells from 3 experiments; ****p < 0.0001; unpaired t-test, two-tailed).

F. Percentage of USH cells with puncta following addition of DMSO or 10 μM CCCP (ns = 0.0594; unpaired t-test, two-tailed).

G. Puncta area in USH cells when treated with 10 μM CCCP (n = 44 puncta from 3 experiments).

H. Number of puncta per DSH cell following treatment with DMSO (n = 75 cells from 4 experiments) or 10 μM CCCP (n = 102 cells from 4 experiments; *p = 0.0278; unpaired t-test, two-tailed).

I. Percentage of DSH cells with puncta following addition of DMSO or 10 μM CCCP (*p = 0.0484; unpaired t-test, two-tailed).

J. Puncta area in DSH cells when treated with 10 μM CCCP (n = 48 puncta from 2 experiments).

For all images, scale bars represent 10 μm. Bar graphs display mean ± standard deviation.