Transgenic Anopheles mosquitoes expressing human PAI-1 impair malaria transmission

Abstract

In mammals, the serine protease plasmin degrades extracellular proteins during blood clot removal, tissue remodeling, and cell migration. The zymogen plasminogen is activated into plasmin by two serine proteases: tissue-type plasminogen activator (tPA) and urokinase-type plasminogen activator (uPA), a process regulated by plasminogen activator inhibitor 1 (PAI-1), a serine protease inhibitor that specifically inhibits tPA and uPA. Plasmodium gametes and sporozoites use tPA and uPA to activate plasminogen and parasite-bound plasmin degrades extracellular matrices, facilitating parasite motility in the mosquito and the mammalian host. Furthermore, inhibition of plasminogen activation by PAI-1 strongly blocks infection in both hosts. To block parasite utilization of plasmin, we engineered Anopheles stephensi transgenic mosquitoes constitutively secreting human PAI-1 (huPAI-1) in the midgut lumen, in the saliva, or both. Mosquitoes expressing huPAI-1 strongly reduced rodent and human Plasmodium parasite transmission to mosquitoes, showing that co-opting plasmin for mosquito infection is a conserved mechanism among Plasmodium species. huPAI-1 expression in saliva induced salivary gland deformation which affects sporozoite invasion and P. berghei transmission to mice, resulting in significant levels of protection from malaria. Targeting the interaction of malaria parasites with the fibrinolytic system using genetically engineered mosquitoes could be developed as an intervention to control malaria transmission.

Combining transgenesis with paratransgenesis to fight malaria

“…That expression of antimalaria effectors in the salivary glands inhibited oocyst development in the midgut (Sg/E; Figure 3A ) is most likely explained by the fact that mosquitoes ingest saliva with the blood meal, in this way incorporating effector proteins into the blood bolus ( Luo et al, 2000 ). A similar phenomenon was also observed in a recent report showing that human PAI-1 expressed in salivary glands was ingested together with the saliva and inhibited oocyst formation ( Pascini et al, 2022 ). In our study, we confirmed the presence of the multi-effector protein, in addition to the AAPP salivary gland protein (control), in the midguts of Sg/E mosquitoes that express the multi-effector protein only in the salivary glands ( Appendix 1—figure 7 ).…”

Activity Sensing of Coagulation and Fibrinolytic Proteases

“…(the latter two not under physiological conditions). [28][29][30][31] SERPINs of fibrinolysis are plasminogen activator inhibitors 1 and 2 (PAI-1/2) and α 2 -antiplasmin, whereas PAI-2 is predominantly intracellular, and its role in the regulation of fibrinolysis is controversial. [32][33][34][35] In order to study and monitor coagulation-related proteases, their respective substrates and specificities need to be explored.…”

“…Fibrinolysis is the process of fibrin clot degradation by plasmin [27] . Activation of plasminogen into plasmin can occur by various activators: tPA, urokinase‐type Plasminogen Activator (uPA, mainly extravascular), bacterial activators, FXIIa, and kallikrein from blood plasma (the latter two not under physiological conditions) [28–31] . SERPINs of fibrinolysis are plasminogen activator inhibitors 1 and 2 (PAI‐1/2) and α 2 ‐antiplasmin, whereas PAI‐2 is predominantly intracellular, and its role in the regulation of fibrinolysis is controversial [32–35] …”

Transgenic approaches in medical entomology: 2022 highlights

“…A recent example of mosquito transgenesis using mosquitoes refractory to Plasmodium by expressing a mosquito restriction factor, has been published by Pascini et al (2022). Following the infectious blood meal, Plasmodium parasites bind plasminogen to their surface in the mosquito midgut lumen.…”