Cell-surface tethered promiscuous biotinylators enable comparative small-scale surface proteomic analysis of human extracellular vesicles and cells.

Autor:	Kirkemo LL; Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, United States., Elledge SK; Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, United States., Yang J; Department of Urology, University of California, San Francisco, San Francisco, United States.; The Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research, University of California, San Francisco, San Francisco, United States., Byrnes JR; Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, United States., Glasgow JE; Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, United States., Blelloch R; Department of Urology, University of California, San Francisco, San Francisco, United States.; The Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research, University of California, San Francisco, San Francisco, United States., Wells JA; Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, United States.; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, United States.
Jazyk:	angličtina
Zdroj:	ELife [Elife] 2022 Mar 08; Vol. 11. Date of Electronic Publication: 2022 Mar 08.
DOI:	10.7554/eLife.73982
Abstrakt:	Characterization of cell surface proteome differences between cancer and healthy cells is a valuable approach for the identification of novel diagnostic and therapeutic targets. However, selective sampling of surface proteins for proteomics requires large samples (>10e6 cells) and long labeling times. These limitations preclude analysis of material-limited biological samples or the capture of rapid surface proteomic changes. Here, we present two labeling approaches to tether exogenous peroxidases (APEX2 and HRP) directly to cells, enabling rapid, small-scale cell surface biotinylation without the need to engineer cells. We used a novel lipidated DNA-tethered APEX2 (DNA-APEX2), which upon addition to cells promoted cell agnostic membrane-proximal labeling. Alternatively, we employed horseradish peroxidase (HRP) fused to the glycan-binding domain of wheat germ agglutinin (WGA-HRP). This approach yielded a rapid and commercially inexpensive means to directly label cells containing common N-Acetylglucosamine (GlcNAc) and sialic acid glycans on their surface. The facile WGA-HRP method permitted high surface coverage of cellular samples and enabled the first comparative surface proteome characterization of cells and cell-derived small extracellular vesicles (EVs), leading to the robust quantification of 953 cell and EV surface annotated proteins. We identified a newly recognized subset of EV-enriched markers, as well as proteins that are uniquely upregulated on Myc oncogene-transformed prostate cancer EVs. These two cell-tethered enzyme surface biotinylation approaches are highly advantageous for rapidly and directly labeling surface proteins across a range of material-limited sample types. Competing Interests: LK, SE, JY, JB, JG, RB, JW No competing interests declared (© 2022, Kirkemo et al.)
Databáze:	MEDLINE
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