Fig. 8.

iRhom2 interacts with proteins involved in vesicle-mediated intracellular transport. Volcano plots of the quantitative comparison of a wild-type iRhom2 vs vector GFP (control) and c mutant iRhom2 (W538S) vs vector control (GFP) co-immunoprecipitations from HEK293 cells based on label-free quantification. Significant regulated proteins are labelled orange (requirements: p-value < 0.01, difference/ratio: > fourfold). All proteins belonging to the GO Group “endoplasmic reticulum to Golgi vesicle-mediated transport” (GO:0006888) and syntaxin 6 (STX6) and syntaxin 7 (STX) as well as the already known main iRhom2 interactors are labelled. The volcano plots were generated using Instant Clue [60]. b, d String images of all proteins belonging to the GO Group “endoplasmic reticulum to Golgi vesicle-mediated transport” (GO:0006888) as well as syntaxin 6 (STX6) and syntaxin 7 (STX7) are shown. All these proteins were found in the quantitative comparison of b wild type iRhom2 vs vector control (GFP) and d mutant iRhom2 (W538S) vs vector control (GFP) co-immunoprecipitations based on label-free quantification. (Requirements: p-value < 0.01, difference/ratio: > fourfold). Of note, SEC22A was not found in the interactome screen but identified by western blot (f). The images were obtained from String v11.0 (string-db.org) [61]. e–g HEK293 cells stably expressing the indicated HA-tagged iRhom constructs were additionally transfected with myc-tagged syntaxin 6 (STX6_myc) (e), syntaxin 10 (STX10_myc) (f) or SEC22a_myc (g). Co-IP experiments with the different iRhom2 constructs as bait were performed and used for western blotting. To probe for the myc-tagged proteins a α-myc antibody was used. Quantitative analysis binding to iRhom2 can be found in figures S10a, b, c. n = 3