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Free keywords:
APEX; RAPIDS; VAPB; biotin; emerin; endoplasmic reticulum (ER); nuclear envelope; nuclear pore; peroxidase; proteomic proximity mapping
Abstract:
VAPB (vesicle-associated membrane protein-associated protein B) is a tail-anchored protein that is present at several contact sites of the endoplasmic reticulum (ER). We now show by immunoelectron microscopy that VAPB also localizes to the inner nuclear membrane (INM). Using a modified APEX2 (enhanced ascorbate peroxidase 2)-approach with rapamycin-dependent targeting of the peroxidase to a protein of interest, we searched for proteins that are in close proximity to VAPB, particularly at the INM. In combination with stable isotope labeling with amino acids in cell culture (SILAC), we confirmed many well-known interaction partners at the level of the ER with a clear distinction between specific and non-specific hits. Furthermore, we identified emerin, TMEM43 and ELYS as potential interaction partners of VAPB at the INM and the nuclear pore complex, respectively.
