Identification of membrane proteins regulated by ADAM15 by SUSPECS proteomics

Calligaris, Matteo, Yang, Chun Y., Bonelli, Simone, Spanò, Donatella Pia, Müller, Stephan A., Lichtenthaler, Stefan F., Troeberg, Linda ORCID: and Scilabra, Simone D. (2023) Identification of membrane proteins regulated by ADAM15 by SUSPECS proteomics. Frontiers in Molecular Biosciences, 10. ISSN 2296-889X

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ADAM15 is a member of the disintegrin-metalloproteinase family of sheddases, which plays a role in several biological processes including cartilage homeostasis. In contrast with well-characterized ADAMs, such as the canonical sheddases ADAM17 and ADAM10, little is known about substrates of ADAM15 or how the enzyme exerts its biological functions. Herein, we used “surface-spanning enrichment with click-sugars (SUSPECS)” proteomics to identify ADAM15 substrates and/or proteins regulated by the proteinase at the cell surface of chondrocyte-like cells. Silencing of ADAM15 by siRNAs significantly altered membrane levels of 13 proteins, all previously not known to be regulated by ADAM15. We used orthogonal techniques to validate ADAM15 effects on 3 of these proteins which have known roles in cartilage homeostasis. This confirmed that ADAM15-silencing increased cell surface levels of the programmed cell death 1 ligand 2 (PDCD1LG2) and reduced cell surface levels of vasorin and the sulfate transporter SLC26A2 through an unknown post-translational mechanism. The increase of PDCD1LG2 by ADAM15 knockdown, a single-pass type I transmembrane protein, suggested it could be a proteinase substrate. However, shed PDCD1LG2 could not be detected even by a data-independent acquisition mass spectrometry, a highly sensitive method for identification and quantification of proteins in complex protein samples, suggesting that ADAM15 regulates PDCD1LG2 membrane levels by a mechanism different from ectodomain shedding.

Item Type: Article
Additional Information: Funding Information: This study was supported by The Kennedy Trust for Rheumatology Research. SFL was supported by the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) under Germany’s Excellence Strategy within the framework of the Munich Cluster for Systems Neurology (EXC 2145 SyNergy—ID 390857198). SDS was supported by Fondazione con il Sud under the Programme “Brains to South 2018”—Project “iRhom2: A new therapeutic target in osteoarthritis” (Grant Agreement No. 2018—PDR—00799) and by “National Biodiversity Future Center—NBFC” (CUP: B73C2100130006) CN_ 00000033 granted by Next Generation EU Programme—PNRR—M4C2—CN5 Spoke 6. Publisher Copyright: Copyright © 2023 Calligaris, Yang, Bonelli, Spanò, Müller, Lichtenthaler, Troeberg and Scilabra.
Uncontrolled Keywords: adam15,adams,chondrocytes,ms,metalloproteases,osteoarthritis,proteomics,specs,biochemistry,molecular biology,biochemistry, genetics and molecular biology (miscellaneous) ,/dk/atira/pure/subjectarea/asjc/1300/1303
Faculty \ School: Faculty of Medicine and Health Sciences > Norwich Medical School
UEA Research Groups: Faculty of Medicine and Health Sciences > Research Centres > Metabolic Health
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Depositing User: LivePure Connector
Date Deposited: 25 Aug 2023 16:30
Last Modified: 19 Oct 2023 03:38
DOI: 10.3389/fmolb.2023.1162504

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