title: Defective tubulin organization and proplatelet formation in murine megakaryocytes lacking Rac1 and Cdc42.
abstract: Blood platelets are anuclear cell fragments that are essential for blood clotting. Platelets are produced by bone marrow megakaryocytes (MKs), which extend protrusions, or so-called proplatelets, into bone marrow sinusoids. Proplatelet formation requires a profound reorganization of the MK actin and tubulin cytoskeleton. Rho GTPases, such as RhoA, Rac1, and Cdc42, are important regulators of cytoskeletal rearrangements in platelets; however, the specific roles of these proteins during platelet production have not been established. Using conditional knockout mice, we show here that Rac1 and Cdc42 possess redundant functions in platelet production and function. In contrast to a single-deficiency of either protein, a double-deficiency of Rac1 and Cdc42 in MKs resulted in macrothrombocytopenia, abnormal platelet morphology, and impaired platelet function. Double-deficient bone marrow MKs matured normally in vivo but displayed highly abnormal morphology and uncontrolled fragmentation. Consistently, a lack of Rac1/Cdc42 virtually abrogated proplatelet formation in vitro. Strikingly, this phenotype was associated with severely defective tubulin organization, whereas actin assembly and structure were barely affected. Together, these results suggest that the combined action of Rac1 and Cdc42 is crucial for platelet production, particularly by regulating microtubule dynamics.
authors.affiliation: University of Würzburg, Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, Deutsche Forschungsgemeinschaft Research Center for Experimental Biomedicine, Würzburg, Germany;
authors.forename: Irina
authors.initials: I
authors.lastname: Pleines
authors.forename: Sebastian
authors.initials: S
authors.lastname: Dütting
authors.forename: Deya
authors.initials: D
authors.lastname: Cherpokova
authors.forename: Anita
authors.initials: A
authors.lastname: Eckly
authors.forename: Imke
authors.initials: I
authors.lastname: Meyer
authors.forename: Martina
authors.initials: M
authors.lastname: Morowski
authors.forename: Georg
authors.initials: G
authors.lastname: Krohne
authors.forename: Harald
authors.initials: H
authors.lastname: Schulze
authors.forename: Christian
authors.initials: C
authors.lastname: Gachet
authors.forename: Najet
authors.initials: N
authors.lastname: Debili
authors.forename: Cord
authors.initials: C
authors.lastname: Brakebusch
authors.forename: Bernhard
authors.initials: B
authors.lastname: Nieswandt
mesh_terms: D000818:Animals; D015153:Blotting, Western; D003599:Cytoskeleton / Q000378:metabolism; D006487:Hemostasis / Q000235:genetics; D055016:Megakaryocyte Progenitor Cells / Q000166:cytology / Q000378:metabolism*; D008533:Megakaryocytes / Q000166:cytology / Q000378:metabolism* / Q000648:ultrastructure; D051379:Mice; D057507:Mice, 129 Strain; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D018613:Microscopy, Confocal; D008855:Microscopy, Electron, Scanning; D046529:Microscopy, Electron, Transmission; D008870:Microtubules / Q000378:metabolism; D011554:Pseudopodia / Q000235:genetics / Q000378:metabolism; D013921:Thrombocytopenia / Q000097:blood / Q000235:genetics / Q000378:metabolism; D013927:Thrombosis / Q000097:blood / Q000235:genetics / Q000378:metabolism; D014404:Tubulin / Q000378:metabolism*; D020764:cdc42 GTP-Binding Protein / Q000172:deficiency / Q000235:genetics*; D020830:rac1 GTP-Binding Protein / Q000172:deficiency / Q000235:genetics*
chemical_list: C107953:RAC1 protein, human; D014404:Tubulin; D020764:cdc42 GTP-Binding Protein; D020830:rac1 GTP-Binding Protein