Fig. 1

Proteolytic coverage map of human IL-38 immunoprecipitated from NHK/38 cells and from normal epidermis. A. NHK/38 cells were lyzed and immunoprecipitated with a polyclonal goat anti-IL-38 antibody (IP anti-IL-38, right lane). Normal goat IgG was used as a negative control to assess the specificity of the immunoprecipitation (IP ctrl IgG, left lane). NHK/38 whole-cell lysates (middle lane) and immunoprecipitated proteins were fractionated by SDS-PAGE and IL-38 was detected by Western blotting. The results are representative of n = 6 independent experiments. B. Normal human epidermis was lyzed and immunoprecipitated with a polyclonal goat anti-IL-38 antibody (IP anti-IL-38, right lane). Normal goat IgG was used as a negative control to assess the specificity of the immunoprecipitation (IP ctrl IgG, middle lane). Epidermis whole-cell lysates (left lane) and immunoprecipitated proteins were fractionated by SDS-PAGE and IL-38 was detected by Western blotting. The results are representative of n = 2 independent experiments. C. The amino acid sequence of human IL-38 is shown. Peptides identified by LC-ESI-MS/MS analysis of IL-38 immunoprecipitated from IL-38 expressing NHK cells (NHK/38; top panel) and normal human epidermis (bottom panel) lysates are highlighted in yellow. The identified peptides cover 32% of the NHK/38 IL-38 protein (48/151 aa, top panel) and 23% of endogenous epidermal IL-38 protein (35/152 aa, bottom panel). The detected N-terminal sequence was CSLPMARYYIIK, corresponding to the IL-38 sequence minus the initiator methionine and thus starting at cysteine 2. Post-translationally modified residues (Table S1), including the acetylated N-terminal cysteine, are highlighted in green