Exposure of plasminogen and a novel plasminogen receptor, Plg-RKT, on activated human and murine platelets

Abstract

Plasminogen activation rates are enhanced by cell surface binding. We have previously demonstrated that exogenous plasminogen binds to phosphatidylserine-exposing and spread platelets. Platelets contain plasminogen in their α-granules but secretion of plasminogen from platelets has not been studied. Recently, a novel transmembrane lysine-dependent plasminogen receptor, Plg-RKT, has been described on macrophages. Here, we analyzed the pool of plasminogen in platelets and examined whether platelets express Plg-RKT. Plasminogen content of the supernatant of resting and collagen/thrombin-stimulated platelets was similar. Pre-treatment with the lysine analogue, εACA, significantly increased platelet-derived plasminogen (0.33 nmol/108 plts vs. 0.08 nmol/108 plts) in the stimulated supernatant, indicating a lysine-dependent mechanism of membrane retention. Lysine-dependent, platelet-derived plasminogen retention on thrombin and convulxin activated human platelets was confirmed by flow cytometry. Platelets initiated fibrinolytic activity in fluorescently labelled plasminogen-deficient clots and in turbidimetric clot lysis assays. A 17 kDa band, consistent with Plg-RKT, was detected in the platelet membrane fraction by Western blotting. Confocal microscopy of stimulated platelets revealed Plg-RKT co-localized with platelet-derived plasminogen on the activated platelet membrane. Plasminogen exposure was significantly attenuated in thrombin and convulxin stimulated platelets from Plg-RKT-/- mice compared to Plg-RKT+/+ littermates. Membrane exposure of Plg-RKT was not dependent on plasminogen, as similar levels of the receptor were detected in plasminogen-/- platelets. These data highlight Plg-RKT as a novel plasminogen receptor in human and murine platelets. We show for the first time that platelet-derived plasminogen is retained on the activated platelet membrane and drives local fibrinolysis, by enhancing cell-surface mediated plasminogen activation.

Successful lactation in Plgrkt-deficient female mice caused by a 1-bp deletion of exon4

“…Plg-R KT is considered to play various pathophysiological roles in vivo , and conventionally developed Plgrkt −/− mice were utilized to prove its roles in inflammatory macrophage recruitment (Miles et al ., 2017), cutaneous wound healing (Ny et al ., 2020), platelet activation (Whyte et al ., 2021) and adipose tissue development (Samad et al ., 2021). It has been considered that Pg activation in the mammary grand tissues is essential for normal lactation due to the phenotypes manifested by conventional Plg −/− (Ploplis et al ., 1995) and Plgrkt −/− (Miles et al ., 2018) mice.…”

Location, location, location: Fibrin, cells, and fibrinolytic factors in thrombi

“…Plg-R KT demonstrates affinity for tPA and is known to co-localize with uPAR on monocytes and macrophages ( 92 ). We have subsequently identified Plg-R KT on platelets and found that it is directly responsible for anchoring plasminogen to the activated platelet membrane ( 93 ). Interestingly, while platelets do not express uPAR, we have shown that the platelet membrane stimulates reciprocal activation of scuPA and plasminogen to their active forms ( 94 ), thereby highlighting the importance of cellular surfaces in regulating profibrinolytic activity.…”

Role of Plasminogen Activation System in Platelet Pathophysiology: Emerging Concepts for Translational Applications

“…More recently, Whyte et al demonstrated the presence of Plg-R kt on the platelet surface. Genetic ablation of Plg-R kt reduces plasminogen binding by roughly 50%, suggesting the central role of this receptor in the retention of plasminogen on the activated platelets [ 47 ].…”