title: Restoration of responsiveness of phospholipase Cγ2-deficient platelets by enforced expression of phospholipase Cγ1.
abstract: Receptor-mediated platelet activation requires phospholipase C (PLC) activity to elevate intracellular calcium and induce actin cytoskeleton reorganization. PLCs are classified into structurally distinct β, γ, δ, ε, ζ, and η isoforms. There are two PLCγ isoforms (PLCγ1, PLCγ2), which are critical for activation by tyrosine kinase-dependent receptors. Platelets express both PLCγ1 and PLCγ2. Although PLCγ2 has been shown to play a dominant role in platelet activation, the extent to which PLCγ1 contributes has not been evaluated. To ascertain the relative contributions of PLCγ1 and PLCγ2 to platelet activation, we generated conditionally PLCγ1-deficient, wild-type (WT), PLCγ2-deficient, and PLCγ1/PLCγ2 double-deficient mice and measured the ability of platelets to respond to different agonists. We found that PLCγ2 deficiency abrogated αIIbβ3-dependent platelet spreading, GPVI-dependent platelet aggregation, and thrombus formation on collagen-coated surfaces under shear conditions, which is dependent on both GPVI and αIIbβ3. Addition of exogenous ADP overcame defective spreading of PLCγ2-deficient platelets on immobilized fibrinogen, suggesting that PLCγ2 is required for granule secretion in response to αIIbβ3 ligation. Consistently, αIIbβ3-mediated release of granule contents was impaired in the absence of PLCγ2. In contrast, PLCγ1-deficient platelets spread and released granule contents normally on fibrinogen, exhibited normal levels of GPVI-dependent aggregation, and formed thrombi normally on collagen-coated surfaces. Interestingly, enforced expression of PLCγ1 fully restored GPVI-dependent aggregation and αIIbβ3-dependent spreading of PLCγ2-deficient platelets. We conclude that platelet activation through GPVI and αIIbβ3 utilizes PLCγ2 because PLCγ1 levels are insufficient to support responsiveness, but that PLCγ1 can restore responsiveness if expressed at levels normally achieved by PLCγ2.
authors: Blood Research Institute, BloodCenter of Wisconsin, Milwaukee, Wisconsin, United States of America; Key Laboratory of Developmental Genes and Human Disease, Ministry of Education, Institute of Life Science, Southeast University, Nanjing, Jiangsu, People's Republic of China.
authors: Yongwei
authors: Y
authors: Zheng
authors: Blood Research Institute, BloodCenter of Wisconsin, Milwaukee, Wisconsin, United States of America.
authors: Tamara
authors: T
authors: Adams
authors: Blood Research Institute, BloodCenter of Wisconsin, Milwaukee, Wisconsin, United States of America.
authors: Huiying
authors: H
authors: Zhi
authors: Blood Research Institute, BloodCenter of Wisconsin, Milwaukee, Wisconsin, United States of America.
authors: Mei
authors: M
authors: Yu
authors: Blood Research Institute, BloodCenter of Wisconsin, Milwaukee, Wisconsin, United States of America.
authors: Renren
authors: R
authors: Wen
authors: Blood Research Institute, BloodCenter of Wisconsin, Milwaukee, Wisconsin, United States of America; Department of Pharmacology and Toxicology, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America; Department of Cell Biology, Neurobiology and Anatomy, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America.
authors: Peter J
authors: PJ
authors: Newman
authors: Blood Research Institute, BloodCenter of Wisconsin, Milwaukee, Wisconsin, United States of America; Department of Microbiology and Molecular Genetics, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America.
authors: Demin
authors: D
authors: Wang
authors: Blood Research Institute, BloodCenter of Wisconsin, Milwaukee, Wisconsin, United States of America; Department of Pharmacology and Toxicology, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America; Department of Microbiology and Molecular Genetics, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America.
authors: Debra K
authors: DK
authors: Newman
mesh_terms: D000818:Animals; D001792:Blood Platelets / Q000378:metabolism*; D003094:Collagen / Q000494:pharmacology; D004789:Enzyme Activation; D015870:Gene Expression; D007527:Isoenzymes; D051379:Mice; D018345:Mice, Knockout; D008822:Mice, Transgenic; D051966:Phospholipase C gamma / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism; D015539:Platelet Activation / Q000235:genetics; D010974:Platelet Aggregation / Q000187:drug effects / Q000235:genetics; D010976:Platelet Count; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000378:metabolism; D013927:Thrombosis / Q000378:metabolism
chemical_list: D007527:Isoenzymes; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D003094:Collagen; D051966:Phospholipase C gamma; C117399:Plcg1 protein, mouse