title: PEAR1 attenuates megakaryopoiesis via control of the PI3K/PTEN pathway.
abstract: Platelet endothelial aggregation receptor-1 (PEAR1) participates in platelet aggregation via sustaining αIIbβ3 activation. To investigate the role of PEAR1 in platelet formation, we monitored and manipulated PEAR1 expression in vitro in differentiating human CD34(+) hematopoietic stem cells and in vivo in zebrafish embryos. PEAR1 expression rose during CD34(+) cell differentiation up to megakaryocyte (MK) maturation. Two different lentiviral short hairpin knockdowns of PEAR1 did not affect erythropoiesis in CD34(+) cells, but increased colony-forming unit MK cell numbers twofold vs control in clonogenic assays, without substantially modifying MK maturation. The PEAR1 knockdown resulted in a twofold reduction of the phosphatase and TENsin homolog (PTEN) phosphatase expression and modulated gene expression of several phosphatidylinositol 3-kinase (PI3K)-Akt and Notch pathway genes. In zebrafish, Pear1 expression increased progressively during the first 3 days of embryo development. Both ATG and splice-blocking PEAR1 morpholinos enhanced thrombopoiesis, without affecting erythropoiesis. Western blots of 3-day-old Pear1 knockdown zebrafish revealed elevated Akt phosphorylation, coupled to transcriptional downregulation of the PTEN isoform Ptena. Neutralization by morpholinos of Ptena, but not of Ptenb, phenocopied the Pear1 zebrafish knockdown and triggered enhanced Akt phosphorylation and thrombocyte formation. In summary, this is the first demonstration that PEAR1 influences the PI3K/PTEN pathway, a critical determinant of Akt phosphorylation, itself controlling megakaryopoiesis and thrombopoiesis.
authors.affiliation: Center for Molecular and Vascular Biology, Department of Cardiovascular Sciences, KU Leuven, Leuven, Belgium.
authors.forename: Alexandre
authors.initials: A
authors.lastname: Kauskot
authors.forename: Christophe
authors.initials: C
authors.lastname: Vandenbriele
authors.forename: Sophie
authors.initials: S
authors.lastname: Louwette
authors.forename: Rik
authors.initials: R
authors.lastname: Gijsbers
authors.forename: Thomas
authors.initials: T
authors.lastname: Tousseyn
authors.forename: Kathleen
authors.initials: K
authors.lastname: Freson
authors.forename: Peter
authors.initials: P
authors.lastname: Verhamme
authors.forename: Marc F
authors.initials: MF
authors.lastname: Hoylaerts
mesh_terms: D000818:Animals; D001792:Blood Platelets / Q000166:cytology / Q000378:metabolism; D015153:Blotting, Western; D002454:Cell Differentiation; D049109:Cell Proliferation; D005434:Flow Cytometry; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000378:metabolism; D006801:Humans; D007124:Immunoenzyme Techniques; D017403:In Situ Hybridization; D008533:Megakaryocytes / Q000166:cytology / Q000378:metabolism; D051059:PTEN Phosphohydrolase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D058539:Phosphatidylinositol 3-Kinase / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D011956:Receptors, Cell Surface / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D034061:Thrombopoiesis / Q000502:physiology*; D015027:Zebrafish / Q000254:growth & development* / Q000378:metabolism
chemical_list: C502809:PEAR1 protein, human; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D011956:Receptors, Cell Surface; D058539:Phosphatidylinositol 3-Kinase; D051057:Proto-Oncogene Proteins c-akt; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human