Comparative membrane proteomics reveals diverse cell regulators concentrated at the nuclear envelope

ABSTRACT: The nuclear envelope (NE) is a subdomain of the ER with a distinctive protein composition that underpins its role in nuclear compartmentalization. To expand an understanding of NE functions, we refined methods to reveal low abundance transmembrane (TM) proteins concentrated at the NE. De facto, these are predicted to have NE-selective functions. We used label-free proteomics to compare isolated NEs to cytoplasmic membranes to identify candidates with substantial NE enrichment. We then examined candidates by immunofluorescence microscopy to quantify their targeting to the NE vs peripheral ER with ectopic expression in cultured cells. Ten candidates from a validation set were found to significantly target to the NE, including oxidoreductases, enzymes for lipid biosynthesis and regulators of cell growth/survival. In a test of functional relevance, we examined one of the NE-concentrated proteins herein identified, the palmitoyltransferase Zdhhc6. We determined that Zdhhc6 modifies the NE oxidoreductase Tmx4 and modulates Tmx4 levels and NE localization, directly supporting NE-related functions for Zdhhc6. Overall, our methodology has revealed a group of previously unrecognized proteins concentrated at the NE as well as additional candidates. Moreover, it provides a framework for uncovering non-abundant components of the NE proteome, and for understanding previously unrecognized NE functions.

INSTRUMENT(S): Orbitrap Fusion Lumos, LTQ Velos

ORGANISM(S): Mus Musculus (ncbitaxon:10090)

SUBMITTER: John R. Yates III  

PROVIDER: MSV000091154 | MassIVE | Wed Jan 25 19:43:00 GMT 2023

REPOSITORIES: MassIVE