sirolimus and emerin

Vesicle-associated membrane protein-associated protein B (VAPB) 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 enhanced ascorbate peroxidase 2 (APEX2) 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 nonspecific hits. Furthermore, we identified emerin, TMEM43, and ELYS as potential interaction partners of VAPB at the INM and the nuclear pore complex, respectively.

Topics: DNA-(Apurinic or Apyrimidinic Site) Lyase; DNA-Binding Proteins; Endonucleases; Endoplasmic Reticulum; HeLa Cells; Humans; Isotope Labeling; Membrane Proteins; Microscopy, Immunoelectron; Multifunctional Enzymes; Nuclear Envelope; Nuclear Proteins; Protein Binding; Protein Interaction Mapping; Protein Interaction Maps; Protein Transport; Proteomics; Sirolimus; Transcription Factors; Vesicular Transport Proteins