Overlapping SETBP1 gain-of-function mutations in Schinzel-Giedion syndrome and hematologic malignancies
Fig 2
Functional analysis of SETBP1 mutations identified in SGS.
A. Fluorescence measurements in live HEK293 cells expressing YFP-tagged SETBP1 variants. (*** p<0.001 versus wild-type and all mutants, ANOVA). All SETBP1 mutations studied displayed a statistically significant difference compared to wild-type and to all other mutations. This graph is representative of 3 independent experiments performed, with 6 technical replicates per experiment. Bars represent the standard error. B. Relative expression of SETBP1 protein variants in live HEK293 cells treated with MG132 proteasome inhibitor or vehicle only. Bars represent the standard error. (*** p<0.001, * p<0.05, NS: not significant, Student’s T test and Mann-Whitney U test). C. ΔΔG values for degron-βTrCP1 interaction for all germline mutations reported in SETBP1 per residue (** p<0.01 D868 versus other residues; ANOVA). D. Immunoblot of whole cell lysates of HEK293 cells expressing FLAG-tagged SETBP1 variants probed with anti-FLAG antibody. E. Immunoblot of whole-cell lysates of fibroblasts probed with anti-SETBP1 antibody. Fibroblasts were derived from two cases of SGS, one carrying the I871T variant and the other carrying the D868N variant, as well as from two unrelated controls. In D and E, blots were stripped and re-probed with anti-β-actin antibody.