Fig. 5: CD99L2 functions as an interactor and activator of CAPN1.
From: Loss-of-function variants in the CAPN1 activator CD99L2 cause X-linked spastic ataxia
a, b Co-immunoprecipitation (myc TRAP) of myc-His6-CD99L2 in 293T cells. Total protein was visualised using SYPRO Ruby. CAPN1, CAPN2, CAPN10, CSS1 and CAST were detected. CD99L2 was visualised using a myc tag antibody. β-actin served as loading control. Grey arrowheads show ubiquitinated (Ub)-CD99L2, white arrowheads unmodified full-length (fl)-proteins, and black arrowheads protein fragments. c Western blot of myc-His6-CD99L2, CAPN1, α-spectrin, CAST and CSS1 in 293T cells. β-actin served as loading control. bdp, CD99L2 breakdown product. d Quantification of full-length und calpain-cleaved proteins detected in (c). SBDP, calpain-cleaved α-spectrin. n = 3 biologically independent experiments. Student’s t-test. e Western blot of CD99L2, CAST, CAPN1, and α-spectrin in 293T cells co-expressing myc-His6-CD99L2 and CAST. β-actin served as loading control. f Quantification of proteins detected in (e). n = 4 biologically independent experiments. One-way ANOVA with Tukey’s post hoc test or Student’s t-test. g Western blot of CD99L2, CAST, CAPN1 and α-spectrin in 293T cells expressing myc-His6-CD99L2 upon calpain inhibitor III (CI III) treatment. β-actin served as loading control. h Quantification of markers detected in (i). n = 4 biologically independent experiments. One-way ANOVA with Tukey’s post hoc test or Student’s t-test. Bars represent mean ± s.e.m. All t-tests are two-tailed. P values are shown in the graphs; ns not significant. Source data are provided as a Source Data file.
