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The tetraspanin Tspan15 is an essential subunit of an ADAM10 scissor complex

Koo, Chek Ziu; Harrison, Neale; Noy, Peter J.; Szyroka, Justyna; Matthews, Alexandra L.; Hsia, Hung En; M�ller, Stephan A.; T�shaus, Johanna; Goulding, Joelle; Willis, Katie; Apicella, Clara; Cragoe, Bethany; Davis, Edward; Keles, Murat; Malinova, Antonia; McFarlane, Thomas A.; Morrison, Philip R.; Nguyen, Hanh T.H.; Sykes, Michael C.; Ahmed, Haroon; Di Maio, Alessandro; Seipold, Lisa; Saftig, Paul; Cull, Eleanor; Pliotas, Christos; Rubinstein, Eric; Poulter, Natalie S.; Briddon, Stephen J.; Holliday, Nicholas D.; Lichtenthaler, Stefan F.; Tomlinson, Michael G.

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Authors

Chek Ziu Koo

Neale Harrison

Peter J. Noy

Justyna Szyroka

Alexandra L. Matthews

Hung En Hsia

Stephan A. M�ller

Johanna T�shaus

Katie Willis

Clara Apicella

Bethany Cragoe

Edward Davis

Murat Keles

Antonia Malinova

Thomas A. McFarlane

Philip R. Morrison

Hanh T.H. Nguyen

Michael C. Sykes

Haroon Ahmed

Alessandro Di Maio

Lisa Seipold

Paul Saftig

Eleanor Cull

Christos Pliotas

Eric Rubinstein

Natalie S. Poulter

Stefan F. Lichtenthaler

Michael G. Tomlinson



Abstract

© 2020 Koo et al. A disintegrin and metalloprotease 10 (ADAM10) is a transmembrane protein essential for embryonic development, and its dysregulation underlies disorders such as cancer, Alzheimer's disease, and inflammation. ADAM10 is a "molecular scissor" that proteolytically cleaves the extracellular region from >100 substrates, including Notch, amyloid precursor protein, cadherins, growth factors, and chemokines. ADAM10 has been recently proposed to function as six distinct scissors with different substrates, depending on its association with one of six regulatory tetraspanins, termed TspanC8s. However, it remains unclear to what degree ADAM10 function critically depends on a TspanC8 partner, and a lack of monoclonal antibodies specific for most TspanC8s has hindered investigation of this question. To address this knowledge gap, here we designed an immunogen to generate the first monoclonal antibodies targeting Tspan15, a model TspanC8. The immunogen was created in an ADAM10-knockout mouse cell line stably overexpressing human Tspan15, because we hypothesized that expression in this cell line would expose epitopes that are normally blocked by ADAM10. Following immunization of mice, this immunogen strategy generated four Tspan15 antibodies. Using these antibodies, we show that endogenous Tspan15 and ADAM10 co-localize on the cell surface, that ADAM10 is the principal Tspan15-interacting protein, that endogenous Tspan15 expression requires ADAM10 in cell lines and primary cells, and that a synthetic ADAM10/Tspan15 fusion protein is a functional scissor. Furthermore, two of the four antibodies impaired ADAM10/Tspan15 activity. These findings suggest that Tspan15 directly interacts with ADAM10 in a functional scissor complex.

Citation

Koo, C. Z., Harrison, N., Noy, P. J., Szyroka, J., Matthews, A. L., Hsia, H. E., …Tomlinson, M. G. (2020). The tetraspanin Tspan15 is an essential subunit of an ADAM10 scissor complex. Journal of Biological Chemistry, 295(36), 12822-12839. https://doi.org/10.1074/jbc.RA120.012601

Journal Article Type Article
Acceptance Date Feb 28, 2020
Online Publication Date Feb 28, 2020
Publication Date Sep 4, 2020
Deposit Date Feb 29, 2020
Publicly Available Date Mar 29, 2024
Journal The 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 295
Issue 36
Pages 12822-12839
DOI https://doi.org/10.1074/jbc.RA120.012601
Keywords tetraspanin, a disintegrin and metalloprotease (ADAM), metalloproteinase, ADAM10, Tspan15, Tspan14, membrane protein, shedding, monoclonal antibody, molecular cell biology
Public URL https://nottingham-repository.worktribe.com/output/4064649
Publisher URL https://www.jbc.org/content/early/2020/02/28/jbc.RA120.012601
Additional Information This research was originally published in the Journal of Biological Chemistry. Chek Ziu Koo1,2*, Neale Harrison1*, Peter J. Noy1*, Justyna Szyroka1, Alexandra L. Matthews1, Hung-En Hsia3, Stephan A. Müller3, Johanna Tüshaus3, Joelle Goulding2,4, Katie Willis1, Clara Apicella1, Bethany Cragoe1, Edward Davis1, Murat Keles1, Antonia Malinova1, Thomas A. McFarlane1, Philip R. Morrison1, Hanh T.H. Nguyen1, Michael C. Sykes1, Haroon Ahmed1, Alessandro Di Maio1, Lisa Seipold5, Paul Saftig5, Eleanor Cull1, Christos Pliotas6, Eric Rubinstein7, Natalie S. Poulter2,8, Stephen J. Briddon2,4, Nicholas D. Holliday4, Stefan F. Lichtenthaler3 and Michael G. Tomlinson1. The tetraspanin Tspan15 is an essential subunit of an ADAM10 scissor complex . J Biol Chem. Year; Vol:pp-pp. © the authors