Fig 1: SLITRK1 physically interacts with L1CAM family proteins.a Coomassie brilliant blue-stained sodium dodecyl-sulfate polyacrylamide gel electrophoresis after immunoprecipitation using anti SLITRK1 or normal rabbit IgG. 200 kDa band (arrow) and 100 kDa band (arrowhead) were excised and subjected for mass-spectrometry analysis (Supplementary Table 3). The blot image was uncropped. b KD, kon, and koff values of SLITRK1 or its variants-binding to Neurofascin, L1CAM, and PTPRd are presented as mean ± SD (Neurofascin and PTPRd, n = 3; L1CAM, n = 6 experiments). c Representative images of cultured cortical neurons overexpressing SLITRK1 and Neurofascin (SL1 + NF). Vec., empty vector (negative control for SLITRK1-expressing plasmid); GFP (control for Neurofascin). d Sholl analysis. Open circle, empty vector + GFP (n = 151 cells); filled circle, SLITRK1 + GFP (n = 117 cells); gray rectangle, SLITRK1 + Neurofascin (n = 103 cells); open rectangle, empty vector + Neurofascin (n = 132 cells). Values are presented as mean ± SEM. e Total branch length. f–h Effects of SLITRK1 on L1CAM-mediated endocytosis of the Sema3a receptor Nrp1. Nrp1/L1CAM-expressing or Nrp1/L1CAM/SLITRK1-expressing COS7 cells were incubated in a medium containing FM4-64 (a fluorescein probe for plasma membrane) with Sema3a or BSA. FM4-64+/Nrp1+/L1CAM+ particles (g) or FM4-64+/Nrp1+/L1CAM+/Sema3a+ particles (h) were counted in each cell. g Vec. (BSA), n = 51; SLITRK1 (BSA), n = 40; Vec. (Sema3a), n = 39; SLITRK1. (Sema3a), n = 45 cells. h Vec., n = 48; WT, n = 48; S330A, n = 51; A444S, n = 48 cells. Values are presented as mean ± SD (e, g, h). *P < 0.05; **P < 0.01; ***P < 0.001 in the Dunnett’s test (b), Steel’s test (e, g, h), or Tukey’s test (d). Scale bar, 50 µm (c, f, low magnification), 10 µm (f, high magnification).