A toolkit for studying cell surface shedding of diverse transmembrane receptors

Abstract
Data availability
Article and author information
Metrics

Abstract

Proteolysis of transmembrane receptors is a critical cellular communication mechanism dysregulated in disease, yet decoding proteolytic regulation mechanisms of hundreds of shed receptors is hindered by difficulties controlling stimuli and unknown fates of cleavage products. Notch proteolytic regulation is a notable exception, where intercellular forces drive exposure of a cryptic protease site within a juxtamembrane proteolytic switch domain to activate transcriptional programs. We created a Synthetic Notch Assay for Proteolytic Switches (SNAPS) that exploits the modularity and unequivocal input/response of Notch proteolysis to screen surface receptors for other putative proteolytic switches. We identify several new proteolytic switches among receptors with structural homology to Notch. We demonstrate SNAPS can detect shedding in chimeras of diverse cell surface receptors, leading to new, testable hypotheses. Finally, we establish the assay can be used to measure modulation of proteolysis by potential therapeutics and offer new mechanistic insights into how DECMA-1 disrupts cell adhesion.

Data availability

All data generated or analysed during this study are included in the manuscript and supporting files. Source data files have been provided for Figures 1 and 3 and 4.

Article and author information

Author details

Amanda N Hayward

Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States

Competing interests
The authors declare that no competing interests exist.
Eric J Aird

Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-4873-042X
Wendy R Gordon

Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States

For correspondence
wrgordon@umn.edu

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0001-7696-5560

Funding

National Institute of General Medical Sciences (R35GM119483)

Wendy R Gordon

Pew Charitable Trusts (Pew Biomedical Scholar)

Wendy R Gordon

National Cancer Institute (U54CA210190)

Eric J Aird

3M (Graduate student fellowship)

Eric J Aird

American Heart Association (Graduate student fellowship)

Amanda N Hayward

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.