title: Protein kinase D2 regulates migration and invasion of U87MG glioblastoma cells in vitro.
abstract: Glioblastoma multiforme (GBM) is the most common malignant brain tumor, which, despite combined modality treatment, reoccurs and is invariably fatal for affected patients. Recently, a member of the serine/threonine protein kinase D (PRKD) family, PRKD2, was shown to be a potent mediator of glioblastoma growth. Here we studied the role of PRKD2 in U87MG glioblastoma cell migration and invasion in response to sphingosine-1-phosphate (S1P), an activator of PRKD2 and a GBM mitogen. Time-lapse microscopy demonstrated that random cell migration was significantly diminished in response to PRKD2 silencing. The pharmacological PRKD family inhibitor CRT0066101 decreased chemotactic migration and invasion across uncoated or matrigel-coated Transwell inserts. Silencing of PRKD2 attenuated migration and invasion of U87MG cells even more effectively. In terms of downstream signaling, CRT0066101 prevented PRKD2 autophosphorylation and inhibited p44/42 MAPK and to a smaller extent p54/46 JNK and p38 MAPK activation. PRKD2 silencing impaired activation of p44/42 MAPK and p54/46 JNK, downregulated nuclear c-Jun protein levels and decreased c-Jun(S73) phosphorylation without affecting the NFκB pathway. Finally, qPCR array analyses revealed that silencing of PRKD2 downregulates mRNA levels of integrin alpha-2 and -4 (ITGA2 and -4), plasminogen activator urokinase (PLAU), plasminogen activator urokinase receptor (PLAUR), and matrix metallopeptidase 1 (MMP1). Findings of the present study identify PRKD2 as a potential target to interfere with glioblastoma cell migration and invasion, two major determinants contributing to recurrence of glioblastoma after multimodality treatment.
authors.affiliation: Institute of Molecular Biology and Biochemistry, Medical University of Graz, Graz, Austria.
authors.forename: Eva
authors.initials: E
authors.lastname: Bernhart
authors.affiliation: Institute of Molecular Biology and Biochemistry, Medical University of Graz, Graz, Austria.
authors.forename: Sabine
authors.initials: S
authors.lastname: Damm
authors.affiliation: Institute of Molecular Biology and Biochemistry, Medical University of Graz, Graz, Austria.
authors.forename: Andrea
authors.initials: A
authors.lastname: Wintersperger
authors.affiliation: Institute of Biophysics, Medical University of Graz, Austria.
authors.forename: Trevor
authors.initials: T
authors.lastname: DeVaney
authors.affiliation: Institute of Pharmaceutical Sciences, Department of Pharmaceutical Technology, Karl-Franzens University, Graz, Austria.
authors.forename: Andreas
authors.initials: A
authors.lastname: Zimmer
authors.affiliation: Cancer Research Technology Ltd, London, UK.
authors.forename: Tony
authors.initials: T
authors.lastname: Raynham
authors.affiliation: Cancer Research Technology Ltd, London, UK.
authors.forename: Christopher
authors.initials: C
authors.lastname: Ireson
authors.affiliation: Institute of Molecular Biology and Biochemistry, Medical University of Graz, Graz, Austria. Electronic address: wolfgang.sattler@medunigraz.at.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Sattler
mesh_terms: D001932:Brain Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects / Q000235:genetics; D004353:Drug Evaluation, Preclinical; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D005909:Glioblastoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D006801:Humans; D008246:Lysophospholipids / Q000494:pharmacology; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000235:genetics; D009361:Neoplasm Invasiveness; D000091283:Protein Kinase D2; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D011743:Pyrimidines / Q000494:pharmacology; D034622:RNA Interference / Q000502:physiology; D034741:RNA, Small Interfering / Q000494:pharmacology; D013110:Sphingosine / Q000031:analogs & derivatives / Q000494:pharmacology
chemical_list: C551536:CRT 0066101; D008246:Lysophospholipids; D000091283:Protein Kinase D2; D047428:Protein Kinase Inhibitors; D011743:Pyrimidines; D034741:RNA, Small Interfering; C060506:sphingosine 1-phosphate; D011494:Protein Kinases; D013110:Sphingosine
keywords: C-Jun; Glioblastoma; Invasion; MAPK; PRKD2; Sphingosine-1-phosphate
references.citation: Expert Rev Neurother. 2012 Apr;12(4):421-35
references.pmid: 22449214
references.citation: EMBO J. 1998 Aug 3;17(15):4404-13
references.pmid: 9687508
references.citation: Nature. 1997 Mar 20;386(6622):296-9
references.pmid: 9069290
references.citation: Brain Tumor Pathol. 2011 Feb;28(1):13-24
references.pmid: 21221826
references.citation: Exp Oncol. 2006 Sep;28(3):225-30
references.pmid: 17080017
references.citation: Int J Cancer. 2010 May 15;126(10):2341-52
references.pmid: 19810093
references.citation: Oncogene. 2012 Nov 1;31(44):4655-66
references.pmid: 22249269
references.citation: J Cell Sci. 2004 Sep 15;117(Pt 20):4619-28
references.pmid: 15371522
references.citation: Mol Cancer Res. 2009 Jan;7(1):23-32
references.pmid: 19147534
references.citation: Nature. 2008 Oct 23;455(7216):1061-8
references.pmid: 18772890
references.citation: N Engl J Med. 2005 Mar 10;352(10):987-96
references.pmid: 15758009
references.citation: Int J Radiat Oncol Biol Phys. 2012 Nov 15;84(4):e515-23
references.pmid: 22901381
references.citation: Appl Immunohistochem Mol Morphol. 2005 Jun;13(2):184-9
references.pmid: 15894933
references.citation: Nat Rev Cancer. 2007 Apr;7(4):281-94
references.pmid: 17384583
references.citation: Nat Rev Mol Cell Biol. 2002 Dec;3(12):932-43
references.pmid: 12461559
references.citation: J Neuropathol Exp Neurol. 2010 Jun;69(6):582-92
references.pmid: 20467333
references.citation: Nucleic Acids Res. 2002 May 1;30(9):e36
references.pmid: 11972351
references.citation: J Cell Physiol. 2011 Apr;226(4):1074-81
references.pmid: 20857418
references.citation: J Clin Oncol. 2008 Dec 1;26(34):5610-7
references.pmid: 18981465
references.citation: Cancer Res. 2008 May 15;68(10):3844-53
references.pmid: 18483269
references.citation: EMBO Rep. 2011 Jul 08;12(8):785-96
references.pmid: 21738220
references.citation: Mol Cancer Res. 2008 Sep;6(9):1469-77
references.pmid: 18819934
references.citation: J Mol Med (Berl). 2009 Apr;87(4):363-74
references.pmid: 19189072
references.citation: Nat Cell Biol. 2009 May;11(5):545-56
references.pmid: 19329994
references.citation: Mol Cancer Ther. 2010 May;9(5):1136-46
references.pmid: 20442301
references.citation: J Clin Invest. 2009 Jun;119(6):1420-8
references.pmid: 19487818
references.citation: J Neurooncol. 2011 Sep;104(3):679-87
references.pmid: 21359852
references.citation: Breast Cancer Res. 2009;11(1):R13
references.pmid: 19243594
references.citation: Neuro Oncol. 2011 Jul;13(7):710-24
references.pmid: 21727210
references.citation: J Biol Chem. 1995 Jul 14;270(28):16483-6
references.pmid: 7622446
references.citation: J Clin Oncol. 2006 Mar 10;24(8):1253-65
references.pmid: 16525180
references.citation: Biochem Biophys Res Commun. 2007 May 4;356(2):361-7
references.pmid: 17359934
references.citation: Proc Natl Acad Sci U S A. 2012 Oct 16;109(42):E2875-84
references.pmid: 23027969
references.citation: Science. 2008 Sep 26;321(5897):1807-12
references.pmid: 18772396
references.citation: BMB Rep. 2011 Oct;44(10):659-64
references.pmid: 22026999
references.citation: Mol Cancer. 2011 Oct 17;10:130
references.pmid: 22004682
references.citation: Proc Natl Acad Sci U S A. 2008 Nov 25;105(47):18378-83
references.pmid: 19011111
references.citation: J Cell Physiol. 2007 Jun;211(3):781-90
references.pmid: 17226786
references.citation: PLoS One. 2012;7(8):e42152
references.pmid: 22879913
references.citation: Nat Cell Biol. 2012 Aug;14(8):777-83
references.pmid: 22854810
references.citation: Oncogene. 2014 Feb 27;33(9):1167-80
references.pmid: 23503467
references.citation: Mol Cancer Res. 2012 Feb;10(2):183-96
references.pmid: 22160930
references.citation: Biochim Biophys Acta. 2009 Jul;1791(7):692-6
references.pmid: 19268560
references.citation: Exp Cell Res. 2005 Mar 10;304(1):175-86
references.pmid: 15707584
references.citation: Curr Biol. 2004 Jan 20;14(2):88-98
references.pmid: 14738729
references.citation: J Cell Sci. 2012 Oct 15;125(Pt 20):4800-11
references.pmid: 22797919
references.citation: Cancer Res. 2004 Dec 15;64(24):8939-44
references.pmid: 15604256
references.citation: Exp Cell Res. 2012 Mar 10;318(5):558-69
references.pmid: 22245102
references.citation: Science. 1997 Jan 17;275(5298):400-2
references.pmid: 8994040
references.citation: PLoS One. 2012;7(6):e38737
references.pmid: 22719931
references.citation: J Biol Chem. 2001 Apr 27;276(17):14443-50
references.pmid: 11278600
references.citation: Nat Rev Cancer. 2010 Jul;10(7):489-503
references.pmid: 20555359
references.citation: BMC Bioinformatics. 2006 Aug 10;7:373
references.pmid: 16901352
references.citation: Cancer Lett. 2003 Sep 10;199(1):53-60
references.pmid: 12963123
references.citation: Nat Rev Cancer. 2010 Jan;10(1):9-22
references.pmid: 20029421
references.citation: J Biol Chem. 2012 May 18;287(21):17065-17076
references.pmid: 22457358
references.citation: Biochim Biophys Acta. 2010 Dec;1806(2):183-92
references.pmid: 20580776
references.citation: Cancer Cell. 2007 May;11(5):447-60
references.pmid: 17482134
references.citation: Biochem Pharmacol. 2010 Dec 15;80(12):1946-54
references.pmid: 20621068
references.citation: J Cell Physiol. 2010 Nov;225(2):472-81
references.pmid: 20458747
references.citation: J Neuropathol Exp Neurol. 2005 Aug;64(8):695-705
references.pmid: 16106218
references.citation: Oncogene. 2003 Dec 4;22(55):8939-47
references.pmid: 14654790
references.citation: Mol Carcinog. 2009 Oct;48(10):910-9
references.pmid: 19326372
references.citation: Cancer Lett. 2002 Jul 26;181(2):195-204
references.pmid: 12175535
references.citation: Cell Signal. 2009 Feb;21(2):253-63
references.pmid: 19038333
references.citation: J Biol Chem. 2009 Jan 9;284(2):799-806
references.pmid: 19001381
references.citation: Biochem J. 2012 Mar 15;442(3):649-59
references.pmid: 22233340
references.citation: PLoS One. 2012;7(8):e43398
references.pmid: 22912867
references.citation: Gut. 2010 Oct;59(10):1316-30
references.pmid: 20732914
references.citation: EMBO J. 2004 Jul 7;23(13):2531-43
references.pmid: 15192707
references.citation: Sci Rep. 2012;2:516
references.pmid: 22816039
references.citation: Cancer Cell. 2003 Jul;4(1):67-79
references.pmid: 12892714
references.citation: Neurosci Res. 2008 Jan;60(1):40-9
references.pmid: 17980449