Fig. 5: Comparison with AP-MS confirmed the dynamic and weak in vivo interactome of STS1. | Nature Communications

Fig. 5: Comparison with AP-MS confirmed the dynamic and weak in vivo interactome of STS1.

From: Spatiotemporal profiling of cytosolic signaling complexes in living cells by selective proximity proteomics

Fig. 5

a Comparison of temporal proteins identified by BP5-based proximity proteomics and FLAG tag-based AP-MS. AP-MS was performed in the same HeLa cells with stably expressed APEX-FLAG-STS1 and EGF stimulation as indicated in Fig. 4a. b Individual temporal curves of the STS1-interacting proteins identified in both APEX2-BP5 proximity proteomics and AP-MS. CBLB, CD2AP, INPPL1, and CSK meet the cutoff only in APEX2-BP5 proximity proteomics or AP-MS (as indicated in brackets), but were manually included into overlap part because of their curve similarity. c Individual temporal curves of the STS1-interacting proteins uniquely significant in AP-MS. Data in b and c are presented as mean values ± s.d. (error bars; n = 3 independent biological experiments). d Validation of CD2AP as an EGF-stimulation-dependent STS1-interacting protein by streptavidin pull-down of stably expressed APEX2-FLAG-CD2AP in HeLa cells and western blots (n = 3 independent biological experiments). Quantification was presented in Supplementary Fig. 12d. e Colocalization of transiently co-transfected GFP-tagged CD2AP and mCherry-tagged STS1 in HeLa cells upon EGF stimulation (n = 3 independent experiments). The scale bars are 10 µm. Results were quantitatively confirmed by line profile analysis shown in Supplementary Fig. 12b. Source data are provided as a Source Data file.

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