Characterising the tetraspanin/ADAM10 ‘molecular scissors’ for the platelet collagen and fibrin receptor GPVI

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Koo, Chek Ziu ORCID: https://orcid.org/0000-0003-3770-2594 (2022). Characterising the tetraspanin/ADAM10 ‘molecular scissors’ for the platelet collagen and fibrin receptor GPVI. University of Birmingham. Ph.D.

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Abstract

ADAM10 is a ubiquitous and essential transmembrane ‘molecular scissor’ that cleaves the ectodomains of >100 different membrane proteins. ADAM10 is regulated by interaction with six tetraspanins (Tspan5, Tspan10, Tspan14, Tspan15, Tspan17 or Tspan33) termed TspanC8s. Importantly, emerging evidence suggests that each TspanC8 promotes cleavage of different ADAM10 substrates, leading to proposal of the ‘six scissors’ hypothesis which suggests that each TspanC8/ADAM10 scissor has a distinct substrate repertoire. The platelet-activating collagen and fibrin receptor GPVI is a promising anti-thrombotic target that is also an ADAM10 substrate. Of the three scissors expressed on human platelets, Tspan15/ADAM10 and Tspan33/ADAM10 can cleave GPVI, whereas Tspan14/ADAM10 cannot. The overarching aim of this project was to examine the molecular basis behind the ‘six scissors’ hypothesis using GPVI as a model substrate. This thesis provided evidence of a novel mechanism where TspanC8s may differentially regulate ADAM10 substrate specificity by controlling the protease’s access to the cut site of a substrate. Tspan15/ADAM10 was identified as the most efficient scissor for GPVI. The extracellular region of Tspan15 was essential for ADAM10 cleavage of GPVI and its cytoplasmic domain appeared to have a minor negative regulatory role. Analysis of non-coding TSPAN15 polymorphisms revealed that high TSPAN15 expression may confer protection against venous thrombosis. ADAM10 and Tspan15 regulated each other’s expression at the protein level, and the cytoplasmic domain of ADAM10 was required for Tspan15 surface expression. Finally, this thesis provided evidence that functional redundancy of Tspan15/ADAM10 with other TspanC8/ADAM10 scissors was minimal, even at high expression levels. These findings may inform the feasibility and the design of future therapeutics that may allow substrate-specific modulation of ADAM10 activity by targeting the regulatory TspanC8 tetraspanin.

Type of Work:

Thesis (Doctorates > Ph.D.)

Award Type:

Doctorates > Ph.D.

Supervisor(s):