The mutant-specific C terminus extension supports the N-domain of CALR for c-MPL binding. (A) A schematic representation of truncated proteins examined in the assay and quantified results obtained from panel B. (B) Co-IP assay for a series of truncated mutant of FLAG-tagged CALR Ins5 (scheme shown in A) and V5-tagged c-MPL (bait) cotransfected into HEK293T cells. Ten percent In and the IP fraction were run as a pair. *Indicates the band from IgG used for immunoprecipitation. Representative data from multiple experiments are presented. Co-IP assays in these 2 panels were performed simultaneously and the data were obtained concurrently. (C) A hypothetical model for CALR Ins5-specific binding to c-MPL. The mutant-specific C terminus extension blocks the P-domain interference and allows the N-domain to bind to c-MPL.