2025/11/17 by Emma M. Webb, Kendal Seymour, Heidi E. Hamm
Biochemistry, Genetics and Molecular Biology · Medicine · #Blood Coagulation and Thrombosis Mechanisms #Platelet Disorders and Treatments #Protease and Inhibitor Mechanisms
paper · doi:10.1055/a-2747-9083
openalex publication_date 2025/11/17 · openalex created_date 2025/11/18 · openalex updated_date 2026/06/11
Abstract Human platelets express protease-activated receptor (PAR) 1 and 4 on the platelet membrane. PAR1 activation is crucial for hemostasis, while PAR4 activation produces a prolonged pro-inflammatory platelet response. Once activated, the PAR receptors cause platelet activation and aggregation. The PARs are activated by proteolytic cleavage of their N-terminus, revealing a new N-terminus called the tethered ligand. This tethered ligand folds back and activates the receptor. PAR1 and PAR4 differ in their tethered ligand sequence as well as their signaling cascades; however, they are both activated by various serine proteases, specifically thrombin. Much is known about PAR4 activation via thrombin activation; however, little is known about activation by other proteases. Thrombin cleavage of PAR4 and subsequent activation leads to G proteins, Gαq and Gα12/13, and β-arrestin signaling. The objective of this study is to characterize trypsin activation of PAR4 on human platelets. Platelets were isolated from the whole blood of healthy human volunteers by centrifugation. Platelets were utilized in flow cytometry assays, calcium mobilization assays, and western blot experiments to characterize trypsin activation of human platelets. Here, we demonstrate that trypsin activates human platelets solely through PAR4. Trypsin activation of PAR4 signaling through both Gαq and Gα12/13 is as effective as α-thrombin activation of PAR4. Thrombin may be the main protease to activate PAR4 under typical conditions; however, under pathological conditions such as acute pancreatitis (AP), trypsin activation of PAR4 may be a main driver of platelet activation and disease progression.