Figure S4. VPS13C KD does not influence... | Rockefeller University Press

Figure S4.

$VPS13C KD does not influence LRRK2 recruitment to lysosomes or its kinase activity. (A–C) Representative immunoblots and relative quantification of LRRK2 and LRRK2-S935 protein levels in LysoIP fractions at baseline condition (N = 4). (D–F) Representative immunoblots and relative quantifications of LRRK2 and LRRK2-S935 protein levels in lysosomal fractions after treatment with CQ (100 μM, 16 h) (N = 4). LRRK2 and LRRK2-S935 protein levels in lysosomal fractions are unchanged at baseline and under lysosomal stress conditions. (G and H) Representative immunoblot and relative protein quantification of Rab10 downstream effector protein EHBP1. (I and J) Representative immunoblot and relative protein quantification of Rab10 phosphatase PPM1H. (K) Representative live-cell confocal images of LAMP1-mGFP (green) in LRRK2 PD mutant (R1441G, lower) and isogenic control (upper) dopaminergic neurons with insets showing smaller lysosomes (yellow arrow) in LRRK2 PD-mutant neurons in comparison to the isogenic control (white arrow) (scale bar: 10 µm, inset: 1 µm). Dashed line represents the outline of the cell. (L) Quantification of average lysosomal size in isogenic control and LRRK2 PD-mutant neurons (N = 3, from n = 11 cells). Data represented as mean ± SEM; unpaired two-tailed t test (B, C, E, F, H, J, and L); ns: not significant (B, C, E, F, H, and J), **P < 0.01 (L). Source data are available for this figure: SourceData FS4.$

VPS13C KD does not influence LRRK2 recruitment to lysosomes or its kinase activity. (A–C) Representative immunoblots and relative quantification of LRRK2 and LRRK2-S935 protein levels in LysoIP fractions at baseline condition (N = 4). (D–F) Representative immunoblots and relative quantifications of LRRK2 and LRRK2-S935 protein levels in lysosomal fractions after treatment with CQ (100 μM, 16 h) (N = 4). LRRK2 and LRRK2-S935 protein levels in lysosomal fractions are unchanged at baseline and under lysosomal stress conditions. (G and H) Representative immunoblot and relative protein quantification of Rab10 downstream effector protein EHBP1. (I and J) Representative immunoblot and relative protein quantification of Rab10 phosphatase PPM1H. (K) Representative live-cell confocal images of LAMP1-mGFP (green) in LRRK2 PD mutant (R1441G, lower) and isogenic control (upper) dopaminergic neurons with insets showing smaller lysosomes (yellow arrow) in LRRK2 PD-mutant neurons in comparison to the isogenic control (white arrow) (scale bar: 10 µm, inset: 1 µm). Dashed line represents the outline of the cell. (L) Quantification of average lysosomal size in isogenic control and LRRK2 PD-mutant neurons (N = 3, from n = 11 cells). Data represented as mean ± SEM; unpaired two-tailed t test (B, C, E, F, H, J, and L); ns: not significant (B, C, E, F, H, and J), **P < 0.01 (L). Source data are available for this figure: SourceData FS4.