Fig. 1

TMEM106B was localized to CHMP2B-positive structures, and the T185, S185, and S134N SNPs of TMEM106B were localized to Rab5- and Rab7-positive endosomes. a Confocal images showing the partial colocalization of endogenous transmembrane protein 106B (TMEM106B) and either charged multivesicular body protein 2B (CHMP2B)^WT or CHMP2B^Intron5 (CHMP2B^In5). Flag-tagged CHMP2B^WT and (b) flag-tagged CHMP2B^In5 were expressed in cultured cortical neurons. Twenty-four hours after transfection, the cells were fixed and stained with anti-flag and anti-TMEM106B antibodies. The arrow indicates the colocalization of TMEM106B and CHMP2B. Scale bar: 10 μm. c Each flag-tagged TMEM106B single-nucleotide polymorphism (SNP; T185, S185, and S134N) was transfected into HEK293T cells. Twenty-four h after transfection, western blot analyses were performed with an anti-Flag or anti-glyceraldehyde 3-phosphate dehydrogenase (GAPDH) antibody. GAPDH was used as a loading control. Each flag-tagged TMEM106B SNP (T185, S185, and S134N) was cotransfected with either green fluorescent protein (GFP)-Rab5 (d) or GFP-Rab7 (e) in cultured cortical neurons. Twenty-four hours after transfection, the neurons were fixed and stained with an anti-flag antibody. f The coefficients of colocalization were measured and analyzed with the ImageJ program (Colocalization Indices). The values are presented as the mean ± standard error of the mean (SEM) of three independent replicates. One-way analysis of variance (ANOVA) followed by Tukey’s multiple comparisons test; *p < 0.05, ***p < 0.001; ns, not significant