UCSF DIABETES, ENDOCRINOLOGY & METABOLISM TRAINING PROGRAM FACULTY RESEARCH SUMMARIES

Mechanisms in Cardiovascular Biology and Disease

The coagulation protease thrombin plays a key role in hemostasis and thrombosis in part by triggering responses in platelets. Protease signaling in other cell types is likely important in inflammation, angiogenesis, and embryonic development. How does a protease behave like a hormone to activate cells? We've characterized a family of protease-activated G protein-coupled receptors (PARs) that provide an answer. Thrombin cleaves PAR1's amino terminal exodomain to unmask a new amino terminus that then serves as a tethered peptide ligand, binding intramolecularly to the body of the receptor to cause transmembrane signaling. Mammals have four PARs. PAR1, PAR3 and PAR4 can be activated by thrombin. PAR2 is activated by trypsin and other proteases with trypsin-like specificity. Together with the coagulation cascade, PARs provide a system that links mechanical information in the form of tissue injury to biochemical information and cellular responses. Current work focuses on:

PAR activation . There is strong evidence that PARs 1, 3 and 4 mediate responses to thrombin in vivo, but it is certainly possible that other proteases served as physiological activators of these receptors. Moreover, although PAR2 can be activated by trypsin and other trypsin-like proteases, physiological activators for PAR2 have not been clearly identified. We are attempting to identify novel PAR activators by biochemical and genomic approaches.

PARs in physiology and disease. We have generated mice deficient in PAR1, PAR2, PAR3, or PAR4. These mice have allowed us to determine which PARs account for responses to thrombin and other proteases in platelets and other specific cell types and to show that PARs in platelets play a key role in hemostasis and thrombosis. Ongoing studies use PAR-deficient mice to define the exact role in thrombin signaling in platelets during the formation of thrombi and to probe the role of PARs in inflammation and in other processes.

Protease signaling in embryonic development . Approximately half of PAR1-deficient mouse embryos die at ~E9.5, apparently from bleeding. Endothelial cell-specific expression of a PAR1 by transgene prevented death of PAR1-deficient embryos. Thus PAR1 signaling in endothelial cells, perhaps in response to coagulation proteases, is important for normal blood vessel development. Efforts to identify the specific proteases and endothelial responses involved are in progress. Other studies are dissecting an apparent role for PARs in the development of the CNS.

G protein-coupled receptor signaling in vivo . PAR1 couples to heterotrimeric G proteins of the Gq, Gi, and G12/13 families. We have generated mice in which G12/13 and Gi signaling can be disrupted in a cell-type specific manner. We are using these mice to probe the roles of specific G protein signaling pathways in hemostasis and thrombosis, blood vessel formation, hematopoiesis and other processes.

Selected References

Griffin, C.T., Srinivasan, Y., Zheng, Y.W.,Huang, W., and Coughlin, S.R. (2001) A role for thrombin receptor signaling in endothelial cells during embryonic development.  Science 293:1666-1670

Camerer, E., Duong, D.N., Hamilton, J.R., and Coughlin, S.R. (2003) Combined deficiency of protease-activated receptor-4 and fibrinogen recapitulates the hemostatic defect but not the embryonic lethality of prothrombin deficiency. Blood 103(1):152-4.

Camerer, E., Qazi, A.A., Duong, D.N., Cornelissen, I., Advincula, R., and Coughlin S.R. (2004) Protease-activated receptors and fibrinogen in hematogenous metastasis. Blood 104(2):397-401.

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