Journal: Science Advances

Article Title: LSD1 enzyme inhibitor TAK-418 unlocks aberrant epigenetic machinery and improves autism symptoms in neurodevelopmental disorder models

doi: 10.1126/sciadv.aba1187

Figure Lengend Snippet: ( A ) Chemical structure of TAK-418. ( B to D ) Effects of 3-day TAK-418 treatment on H3K4me2 (B) and H3K9me2 (C) levels at the Ucp2 gene and Ucp2 mRNA levels (D) in primary cultured rat neurons. N = 3 to 4. One-tailed parametric Williams’ test versus DMSO-treated groups, * P < 0.025, ** P < 0.005, and *** P < 0.0005. ( E ) Immunoprecipitation (IP) analyses of the interaction between LSD1 and GFI1B in TF-1a cells. GFI1B/LSD1 values are obtained from densitometry data of GFI1B bands normalized by LSD1 bands (lane 9 = 100%). IgG, immunoglobulin G; WB, Western blotting. ( F ) Superimposed structure of formylated FAD (stick model in white) in human recombinant LSD1 generated after TAK-418 treatment and N-terminal GFI1B peptide (PRSFLV, stick model in yellow) in the LSD1 complex. The formyl-FAD adduct is compact in the active site of LSD1 and has minimal impact on the interaction between LSD1 and GFI1B. ( G ) Dose-dependent change in LSD1 enzyme activity in the rat cortex 2 hours after administration of TAK-418. N = 3 to 6. One-tailed Shirley-Williams test versus vehicle-treated group, ** P < 0.005. Data are shown as means + SD.

Article Snippet: LSD1 and GFI1B proteins bound to the sepharose beads were analyzed by Western blotting using NuPAGE LDS sample buffer (NP0007, Invitrogen), NuPAGE Protein Analysis System with NuPAGE Bis-Tris Gels (NP0323, Invitrogen), NuPAGE MOPS SDS Running Buffer (NP0001, Invitrogen), polyvinylidene difluoride membrane (LC20002, Invitrogen), NuPAGE transfer buffer (NP0006-1, Invitrogen), anti-LSD1 antibody (2139, Cell Signaling Technology), and anti-GFI1B antibody (sc-28356, Santa Cruz).

Techniques: Cell Culture, One-tailed Test, Immunoprecipitation, Western Blot, Recombinant, Generated, Activity Assay