title: A time frame permissive for Protein Kinase D2 activity to direct angiogenesis in mouse embryonic stem cells.
abstract: The protein kinase D isoenzymes PKD1/2/3 are prominent downstream targets of PKCs (Protein Kinase Cs) and phospholipase D in various biological systems. Recently, we identified PKD isoforms as novel mediators of tumour cell-endothelial cell communication, tumour cell motility and metastasis. Although PKD isoforms have been implicated in physiological/tumour angiogenesis, a role of PKDs during embryonic development, vasculogenesis and angiogenesis still remains elusive. We investigated the role of PKDs in germ layer segregation and subsequent vasculogenesis and angiogenesis using mouse embryonic stem cells (ESCs). We show that mouse ESCs predominantly express PKD2 followed by PKD3 while PKD1 displays negligible levels. Furthermore, we demonstrate that PKD2 is specifically phosphorylated/activated at the time of germ layer segregation. Time-restricted PKD2-activation limits mesendoderm formation and subsequent cardiovasculogenesis during early differentiation while leading to branching angiogenesis during late differentiation. In line, PKD2 loss-of-function analyses showed induction of mesendodermal differentiation in expense of the neuroectodermal germ layer. Our in vivo findings demonstrate that embryoid bodies transplanted on chicken chorioallantoic membrane induced an angiogenic response indicating that timed overexpression of PKD2 from day 4 onwards leads to augmented angiogenesis in differentiating ESCs. Taken together, our results describe novel and time-dependent facets of PKD2 during early cell fate determination.
authors.affiliation: Department of Internal Medicine I, Ulm University, Ulm, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Müller
authors.affiliation: Department of Internal Medicine I, Ulm University, Ulm, Germany.
authors.forename: Jana
authors.initials: J
authors.lastname: Schröer
authors.affiliation: Department of Internal Medicine I, Ulm University, Ulm, Germany.
authors.forename: Ninel
authors.initials: N
authors.lastname: Azoitei
authors.affiliation: Department of Internal Medicine I, Ulm University, Ulm, Germany.
authors.forename: Tim
authors.initials: T
authors.lastname: Eiseler
authors.affiliation: Department of Internal Medicine I, Ulm University, Ulm, Germany.
authors.forename: Wendy
authors.initials: W
authors.lastname: Bergmann
authors.affiliation: Department of Internal Medicine I, Ulm University, Ulm, Germany.
authors.forename: Ralf
authors.initials: R
authors.lastname: Köhntop
authors.affiliation: Department of Cell Biology, Institute for Biomedical Engineering, RWTH Aachen University Medical School, Aachen, Germany.
authors.forename: Qiong
authors.initials: Q
authors.lastname: Lin
authors.affiliation: IZKF Computational Biology Research Group, RWTH Aachen University Medical School, Aachen, Germany.
authors.forename: Ivan G
authors.initials: IG
authors.lastname: Costa
authors.affiliation: Department of Cell Biology, Institute for Biomedical Engineering, RWTH Aachen University Medical School, Aachen, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Zenke
authors.affiliation: Department of Urology, Ulm University, Ulm, Germany.
authors.forename: Felicitas
authors.initials: F
authors.lastname: Genze
authors.affiliation: Department of Internal Medicine I, Ulm University, Ulm, Germany.
authors.forename: Clair
authors.initials: C
authors.lastname: Weidgang
authors.affiliation: Department of Internal Medicine I, Ulm University, Ulm, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Seufferlein
authors.affiliation: Institute of Neuroanatomy, Eberhard Karls University Tuebingen, Tuebingen, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Liebau
authors.affiliation: Department of Internal Medicine I, Ulm University, Ulm, Germany.
authors.forename: Alexander
authors.initials: A
authors.lastname: Kleger
mesh_terms: D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D002645:Chickens; D049033:Chorioallantoic Membrane / Q000098:blood supply; D004318:Doxycycline / Q000494:pharmacology; D058732:Embryoid Bodies / Q000166:cytology / Q000637:transplantation; D055879:Gene Knock-In Techniques; D007150:Immunohistochemistry; D057026:Induced Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D000066450:Mouse Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism*; D009389:Neovascularization, Pathologic; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism; D000091283:Protein Kinase D2; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction
chemical_list: D000091283:Protein Kinase D2; D011494:Protein Kinases; C119190:protein kinase C nu; C089212:protein kinase D; D011493:Protein Kinase C; D004318:Doxycycline