A new panel of epitope mapped monoclonal antibodies recognising the prototypical tetraspanin CD81

Abstract

Background: Tetraspanins are small transmembrane proteins, found in all higher eukaryotes, that compartmentalize cellular membranes through interactions with partner proteins. CD81 is a prototypical tetraspanin and contributes to numerous physiological and pathological processes, including acting as a critical entry receptor for hepatitis C virus (HCV). Antibody engagement of tetraspanins can induce a variety of effects, including actin cytoskeletal rearrangements, activation of MAPK-ERK signaling and cell migration. However, the epitope specificity of most anti-tetraspanin antibodies is not known, limiting mechanistic interpretation of these studies. Methods: We generated a panel of monoclonal antibodies (mAbs) specific for CD81 second extracellular domain (EC2) and performed detailed epitope mapping with a panel of CD81 mutants. All mAbs were screened for their ability to inhibit HCV infection and E2-CD81 association. Nanoscale distribution of cell surface CD81 was investigated by scanning electron microscopy. Results: The antibodies were classified in two epitope groups targeting opposing sides of EC2. We observed a wide range of anti-HCV potencies that were independent of their epitope grouping, but associated with their relative affinity for cell-surface expressed CD81. Scanning electron microscopy identified at least two populations of CD81; monodisperse and higher-order assemblies, consistent with tetraspanin-enriched microdomains. Conclusions: These novel antibodies provide well-characterised tools to investigate CD81 function, including HCV entry, and have the potential to provide insights into tetraspanin biology in general.

Publication DOI: https://doi.org/10.12688/wellcomeopenres.12058.1
Divisions: College of Health & Life Sciences
College of Health & Life Sciences > School of Biosciences > Cellular and Molecular Biomedicine
College of Health & Life Sciences > School of Biosciences
Additional Information: Funding: BBSRC (BB/N007417/1); EC (LSHG-CT-2004-504601 and FP7 F3-2012-305578); MRC (MC_UU_12018/1 and G1100247); Wellcome Trust (200838); and Royal Society (107653).
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Related URLs: https://www.sco ... a19ec1cc9a66a1c (Scopus URL)
https://wellcom ... rticles/2-82/v1 (Publisher URL)
PURE Output Type: Article
Published Date: 2017-09-07
Published Online Date: 2017-09-07
Accepted Date: 2017-09-07
Authors: Grove, Joe
Hu, Ke
Farquhar, Michelle J.
Goodall, Margaret
Walker, Lucas
Jamshad, Mohammed
Drummer, Heidi E.
Bill, Roslyn M. (ORCID Profile 0000-0003-1331-0852)
Balfe, Peter
McKeating, Jane A.

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Version: Accepted Version

License: Creative Commons Attribution

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