Sterile20 Kinases TNIK Positively Regulates Platelet Function And Thrombus Formation

Platelet plays an important role in hemostasis and thrombosis.Impaired or excessive activation of platelets leads to increased possibilities of hemorrhage or thrombotic disorders,respectively.The investigation of the potential mechanism and related molecular interaction of platelet proteins will have potential to provide new therapeutic targets and develope possible treatment for the related diseases.As a key signaling network during platelet activation,MAPKs are involved in platelet functional regulation including adhesion,secretion and aggregation,and eventually thrombus formation.MAPK activation causes a sequential activated triple-kinase module,namely MAP3K phosphorylation,MAP2K phosphorylation and then MAPK activation.However,whether MAPKs selectively and specifically regulate platelet remains unclear.Furthermore,sterile-20 kinases,a superfamily of MAP4K,play a critical role in thrombosis formation as well.According to the structure,sterile-20 kinases can be divided into two isoforms:the p21-activated kinases(PAKs)and germinal center kinases(GCKs).Among them,MINK1 and TNIK are both belong to GCKIV subfamily.It has been reported that MINK1 act as a positive regulation for thrombosis,while the role of TNIK,which related to cell proliferation and neurodevelopment,remains unknown in hemostasis and thrombosis.Considering the homologue of TNIK and MINK1,we thus hypothesize that TNIK may participate in platelet activation,resulting to thrombus production via MAPKs.Using TNIK-specific deficient(TNIK-/-)mice,we studied the role of TNIK in platelet function and thrombus formation.The results can be summarized as follows:(1)In ferric chloride induced mesenteric arteriolar thrombosis model,TNIK-/-mice showed a significant increase of vessel occlusion time,almost twice as much as wild-type(WT)mice.(2)In the platelet functional test,TNIK-/-platelets displayed a remarkably impaired aggregation in response to low but not high concentrations of thrombin and collagen.Their a-granule secretion,as measured by P-Selectin expression,is normal.TNIK-/-platelets also showed a significantly decreased spreading area on fibrinogen,indicating it is also involved in platelet outside-in signaling.(3)Importantly,TNIK-/-platelets showed a significantly decreased ADP release stimulated by either low or high concentration of agonists.When ADP was hydrolyzed by Apyrase,the aggregation and spreading differences between TNIK-/-and WT platelets were abolished.Exogenous ADP application can rescue the spreading defects of TNIK-/-platelets.(4)TNIK-deficient lead to the decrease of p38 and AKT Serine 308/Threonin473 phosphorylation.Taken together,TNIK is a novel molecular in platelet activation,hemostasis and thrombosis.TNIK plays a critical role in controlling platelet dense granule release.possibly through the regulation of p38 and AKT.