Spatially restricted proteomics reveals dynamic changes in the caveolin-1 interactome in response to increased membrane tension

Here, we used APEX2 proximity labelling technique coupled with mass spectrometry to define Cveolin-1 interactome at steady state and after hypo-osmotic shock. We created a stable cell line expressing Caveolin1-APEX2-EGFP in human retinal pigmented epithelial cells (RPE-1). Hypo osmotic shock was performed accordingly to Sinha et al, 2011. Briefly, cells were incubated in 30mOsm medium for 5 minutes (Hypo condition). To analyse the ability to recover, after the osmotic shock cells were allowed to recover in isotonic medium for 30 minutes (REC condition). Cells grown in iso-tonic medium were used as normal non treated condition (NT). RPE-1 cells expressing APEX2-EGFP in the cytoplasm (NES-APEX2-EGFP) was used as control to exclude highly abundant cytosolic proteins.