Figure S1.
Quantification of Cry2WT-mCherry-Tau (CWT 1–441) distribution in SH SY5Y before and after blue-light activation and Tau dissociates from MTs after 1,6-HD treatment in HeLa cells.(A) Cry2WT-mCherry-Tau (CWT 1–441) distribution in SH SY5Y before activation (green dots), after activation (orange dots), and change by activation (blue dots) were quantified using pixel CV (y axis) for each cell (x axis). The CV of the Tau signal after blue-light activation is significantly higher than before, n = 23, P = 7.0 × 10−7, paired t test. (B–D) Full-length Tau 1–441-EGFP is sensitive to 1,6-HD but not to 2,5-HD. (B) Protein schematic diagram of Tau 1–441-EGFP. (C) Tau 1–441-EGFP expressed in HeLa cells was treated with 1,6-HD, which resulted in the loss of a MT-binding pattern to a diffuse pattern. Selected images at 0 min, 1.5 min, and 3 min are presented. (D) Tau 1–441-EGFP was treated with 2,5-HD in HeLa cell; selected images at 0 min, 3 min, and extended time to 20 min are presented. Tau binds to MT even after extended treatment of 2,5-HD. (E and F) Tau truncations containing MTBD and the extreme C terminus remain sensitive to 1,6-HD treatment, and MT was not affected. (E) Protein schematic diagram of Tau 256–441-EGFP. (F) Tau 256–441-EGFP (green) and mCherry-α-tubulin (red) were overexpressed in HeLa cells, treated with 1,6-HD. Montage of selected images at 0 min and 3 min treatment are presented. From left to right, mCherry-α-tubulin, Tau 256–441-EGFP, and the merge. While truncated Tau became diffuse after 3 min, mCherry-α-tubulin still maintained the MT pattern. Note that there is a nuclear population of EGFP signal for Tau 256–441-EGFP overexpression, which is commonly seen for EGFP fused with proteins of low molecular weight. (G) Quantification of Tau distribution upon 1,6-HD or 2,5-HD treatment based upon CV. Distribution of Tau 1–441-EGFP treated with 1,6-HD (black), Tau 1–441-EGFP treated with 2,5-HD (orange), Tau 256–441-EGFP treated with 1,6-HD (gray), and mCherry-a-tubulin treated with 1,6-HD (yellow) was evaluated and plotted over the time. ID, identification; t, time.

Quantification of Cry2WT-mCherry-Tau (CWT 1–441) distribution in SH SY5Y before and after blue-light activation and Tau dissociates from MTs after 1,6-HD treatment in HeLa cells. (A) Cry2WT-mCherry-Tau (CWT 1–441) distribution in SH SY5Y before activation (green dots), after activation (orange dots), and change by activation (blue dots) were quantified using pixel CV (y axis) for each cell (x axis). The CV of the Tau signal after blue-light activation is significantly higher than before, n = 23, P = 7.0 × 10−7, paired t test. (B–D) Full-length Tau 1–441-EGFP is sensitive to 1,6-HD but not to 2,5-HD. (B) Protein schematic diagram of Tau 1–441-EGFP. (C) Tau 1–441-EGFP expressed in HeLa cells was treated with 1,6-HD, which resulted in the loss of a MT-binding pattern to a diffuse pattern. Selected images at 0 min, 1.5 min, and 3 min are presented. (D) Tau 1–441-EGFP was treated with 2,5-HD in HeLa cell; selected images at 0 min, 3 min, and extended time to 20 min are presented. Tau binds to MT even after extended treatment of 2,5-HD. (E and F) Tau truncations containing MTBD and the extreme C terminus remain sensitive to 1,6-HD treatment, and MT was not affected. (E) Protein schematic diagram of Tau 256–441-EGFP. (F) Tau 256–441-EGFP (green) and mCherry-α-tubulin (red) were overexpressed in HeLa cells, treated with 1,6-HD. Montage of selected images at 0 min and 3 min treatment are presented. From left to right, mCherry-α-tubulin, Tau 256–441-EGFP, and the merge. While truncated Tau became diffuse after 3 min, mCherry-α-tubulin still maintained the MT pattern. Note that there is a nuclear population of EGFP signal for Tau 256–441-EGFP overexpression, which is commonly seen for EGFP fused with proteins of low molecular weight. (G) Quantification of Tau distribution upon 1,6-HD or 2,5-HD treatment based upon CV. Distribution of Tau 1–441-EGFP treated with 1,6-HD (black), Tau 1–441-EGFP treated with 2,5-HD (orange), Tau 256–441-EGFP treated with 1,6-HD (gray), and mCherry-a-tubulin treated with 1,6-HD (yellow) was evaluated and plotted over the time. ID, identification; t, time.

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