Fig 2: CMTM4 mediates IL-17RC glycosylation.a, Immunoblotting analysis of samples isolated via immunoprecipitation with IL-17RC antibody or control isotype antibody from lysates of wild-type or Cmtm4-/- ST2 cells. b,c, Immunoblotting analysis of samples isolated via immunoprecipitation with IL-17RC antibody from lysates of wild-type or several Cmtm4-/- ST2 cell lines (b) or Cmtm4-/- ST2 cells expressing Myc-CMTM4 or EV (c). d, Immunoblotting analysis of samples isolated via immunoprecipitation with IL-17RC antibody from lysates of wild-type or Cmtm4-/- ST2 cells treated or not with proteasome inhibitor bortezomib (10 nM) for 24 h. e, Immunoblotting analysis of samples isolated by Flag immunoprecipitation from lysates of Cmtm4-/- ST2 cells transduced with expression vectors coding for indicated Myc-tagged members of CMTM family together with the expression vector coding for SF-IL-17RC. f-j, Immunoblotting analysis of samples isolated by Flag immunoprecipitation from lysates of wild-type or Cmtm4-/- ST2 cells expressing SF-IL-17RA (f), SF-IL-17RB (g), SF-IL-17RD (h), SF-IL-17RE (i), or SF-TNFR1 (j) and treated or not with PNGase F. *, unspecific band. Data are representative of four (a), three (b–d), or two (e–j) independent experiments. Source data

Fig 3: IL-17RC glycosylation is required for trafficking to the plasma membrane.a, Immunoblotting analysis of lysates of ST2 cells transduced with expression vectors coding for SF-IL-17RC harboring indicated mutations of the putative glycosylation sites. b, Flow cytometry analysis of IL-17RC surface expression in cells expressing indicated SF-IL-17RC constructs. c, Immunoblotting analysis of samples isolated through Flag IP from lysates of ST2 cells expressing indicated SF-IL-17RC constructs. d–i, Immunoblotting (d,g), flow cytometry of surface IL-17RC (e,h), and fluorescence microscopy (f,i) in Cmtm4-/- ST2 cells expressing IL-17RC-EGFP only (d,e,f) or together with CMTM4-mCherry (g,h,i). Scale bar, 5 µm. Data are representative of three (a–c,f,i) or two (d,g) independent experiments. Flow cytometry data show mean + s.e.m. from three (b) or five (e,h) independent experiments. Two-tailed Mann–Whitney test (e,h) or two-tailed unpaired t-test (b).Source data

Fig 4: CMTM4 is constitutively associated with IL-17RC transmembrane domain.a, MS analysis of IL-17RSC isolated via tandem affinity purification from lysates of ST2 cells that were stimulated with SF-IL-17A (500 ng ml–1) for 15 min. Proteins identified in stimulated samples, but not in control samples in which SF-IL-17A was added after lysis, are listed. Relative quantification is based on iBAQ values from 13 experiments. b, Immunoblotting analysis of samples isolated by Flag IP from lysates of ST2 cells transduced with expression vectors coding for SF-IL-17RA, SF-IL-17RC or EV. c,d, Immunoblotting analysis of samples isolated by Flag IP from lysates of ST2 cells expressing EV or SF-IL-17RC (c) or SF-IL-17RA (d) that were stimulated with IL-17A (500 ng ml–1) for 15 min or left unstimulated. *, unspecific band. e, Immunoblotting analysis of samples isolated through IP with IL-17RC antibody or isotype control antibody from the lysates of ST2 cells stimulated with IL-17A (500 ng ml–1) for 15 min or left unstimulated. f, MS analysis of proteins isolated via tandem affinity purification from the lysates of ST2 cells expressing SF-IL-17RC. Relative quantification is based on iBAQ values from three experiments. Only proteins that were not detected in control samples isolated from ST2 cells expressing SF-EGFP and that had an average iBAQ value >2 × 106 are listed. g, Mean + s.e.m. of iBAQ ratio between CMTM4 and IL-17RC in IL-17RSC calculated for individual MS experiments shown in a. h, Immunoblotting analysis of samples isolated by Flag IP from lysates of Drosophila S2 cells transduced with expression vectors coding for SF-IL-17RA, SF-IL-17RC or EV together with the expression vector coding for Myc-CMTM4. i, Immunoblotting analysis of samples isolated through Flag IP from lysates of ST2 cells transduced with expression vectors coding for SF-IL-17RA, SF-IL-17RC or indicated chimeric proteins. j, Immunoblotting analysis of samples isolated by Flag IP from lysates of ST2 cells expressing CMTM4-Myc, CMTM6-Myc or indicated chimeric proteins together with SF-IL-17RC. Data are representative of two (d,i,j), three (c,h) or four (b,e) independent experiments.Source data

Fig 5: CMTM4 mediates IL-17RC glycosylation and plasma membrane localization.a,b, Immunoblotting analysis of CMTM4 and actin and flow cytometry analysis of IL-17RC surface expression in wild-type and Cmtm4-/- ST2 cells (a) or Cmtm4-/- ST2 cells transduced with expression vectors coding for Myc-CMTM4 or EV (b). c, Immunoblotting analysis of samples isolated via IP with an IL-17RC antibody from lysates of wild-type and Cmtm4-/- ST2 cells stimulated with IL-17A (500 ng ml–1) for 15 min or left unstimulated. d–f, Immunoblotting analysis of samples isolated through Flag IP from lysates of wild-type and Cmtm4-/- ST2 cells expressing SF-IL-17RC that were treated with O-glycosidase (d), PNGase F (d, e) or Endo H (d, f) as indicated. Data are representative of two (c–f) independent experiments. Flow cytometry data are represented as mean + s.e.m. from six (a) or five (b) independent experiments. Two-tailed Mann–Whitney test. MFI, median fluorescence intensity.Source data