Proteomic mapping by rapamycin-dependent targeting of APEX2 identifies binding partners of VAPB at the inner nuclear membrane

James, Christina; Müller, Marret; Goldberg, Martin W.; Lenz, Christof; Urlaub, Henning; Kehlenbach, Ralph H.

doi:10.1074/jbc.ra118.007283

Proteomic mapping by rapamycin-dependent targeting of APEX2 identifies binding partners of VAPB at the inner nuclear membrane

James, Christina; Müller, Marret; Goldberg, Martin W.; Lenz, Christof; Urlaub, Henning; Kehlenbach, Ralph H.

Authors

Christina James

Marret Müller

Professor Martin Goldberg m.w.goldberg@durham.ac.uk
Professor

Christof Lenz

Henning Urlaub

Ralph H. Kehlenbach

Abstract

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.

Citation

James, C., Müller, M., Goldberg, M. W., Lenz, C., Urlaub, H., & Kehlenbach, R. H. (2019). Proteomic mapping by rapamycin-dependent targeting of APEX2 identifies binding partners of VAPB at the inner nuclear membrane. Journal of Biological Chemistry, 294(44), 16241-16254. https://doi.org/10.1074/jbc.ra118.007283

Journal Article Type	Article
Online Publication Date	Sep 13, 2019
Publication Date	Nov 1, 2019
Deposit Date	Sep 25, 2019
Publicly Available Date	Sep 13, 2020
Journal	Journal of Biological Chemistry
Print ISSN	0021-9258
Electronic ISSN	1083-351X
Publisher	American Society for Biochemistry and Molecular Biology
Peer Reviewed	Peer Reviewed
Volume	294
Issue	44
Pages	16241-16254
DOI	https://doi.org/10.1074/jbc.ra118.007283

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Copyright Statement
This research was originally published in James, Christina, Müller, Marret, Goldberg, Martin W., Lenz, Christof, Urlaub, Henning & Kehlenbach, Ralph H. (2019). Proteomic mapping by rapamycin-dependent targeting of APEX2 identifies binding partners of VAPB at the inner nuclear membrane. Journal of Biological Chemistry 294(44): 16241-16254 © the American Society for Biochemistry and Molecular Biology.