title: Cerebral cavernous malformations arise from endothelial gain of MEKK3-KLF2/4 signalling.
abstract: Cerebral cavernous malformations (CCMs) are common inherited and sporadic vascular malformations that cause strokes and seizures in younger individuals. CCMs arise from endothelial cell loss of KRIT1, CCM2 or PDCD10, non-homologous proteins that form an adaptor complex. How disruption of the CCM complex results in disease remains controversial, with numerous signalling pathways (including Rho, SMAD and Wnt/β-catenin) and processes such as endothelial-mesenchymal transition (EndMT) proposed to have causal roles. CCM2 binds to MEKK3 (refs 7, 8, 9, 10, 11), and we have recently shown that CCM complex regulation of MEKK3 is essential during vertebrate heart development. Here we investigate this mechanism in CCM disease pathogenesis. Using a neonatal mouse model of CCM disease, we show that expression of the MEKK3 target genes Klf2 and Klf4, as well as Rho and ADAMTS protease activity, are increased in the endothelial cells of early CCM lesions. By contrast, we find no evidence of EndMT or increased SMAD or Wnt signalling during early CCM formation. Endothelial-specific loss of Map3k3 (also known as Mekk3), Klf2 or Klf4 markedly prevents lesion formation, reverses the increase in Rho activity, and rescues lethality. Consistent with these findings in mice, we show that endothelial expression of KLF2 and KLF4 is increased in human familial and sporadic CCM lesions, and that a disease-causing human CCM2 mutation abrogates the MEKK3 interaction without affecting CCM complex formation. These studies identify gain of MEKK3 signalling and KLF2/4 function as causal mechanisms for CCM pathogenesis that may be targeted to develop new CCM therapeutics.
authors.affiliation: Department of Medicine and Cardiovascular Institute, University of Pennsylvania, 3400 Civic Center Blvd, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Zinan
authors.initials: Z
authors.lastname: Zhou
authors.affiliation: Department of Medicine and Cardiovascular Institute, University of Pennsylvania, 3400 Civic Center Blvd, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Alan T
authors.initials: AT
authors.lastname: Tang
authors.affiliation: Laboratory of Cardiovascular Signaling, Centenary Institute, Sydney, New South Wales 2050, Australia.
authors.forename: Weng-Yew
authors.initials: WY
authors.lastname: Wong
authors.affiliation: Department of Medicine and Cardiovascular Institute, University of Pennsylvania, 3400 Civic Center Blvd, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Sharika
authors.initials: S
authors.lastname: Bamezai
authors.affiliation: Department of Medicine and Cardiovascular Institute, University of Pennsylvania, 3400 Civic Center Blvd, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Lauren M
authors.initials: LM
authors.lastname: Goddard
authors.affiliation: Neurovascular Surgery Program, Section of Neurosurgery, Department of Surgery, The University of Chicago Medicine and Biological Sciences, Chicago, Illinois 60637, USA.
authors.forename: Robert
authors.initials: R
authors.lastname: Shenkar
authors.affiliation: Department of Medicine and Cardiovascular Institute, University of Pennsylvania, 3400 Civic Center Blvd, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Su
authors.initials: S
authors.lastname: Zhou
authors.affiliation: Department of Medicine and Cardiovascular Institute, University of Pennsylvania, 3400 Civic Center Blvd, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Jisheng
authors.initials: J
authors.lastname: Yang
authors.affiliation: Department of Radiology, University of Pennsylvania Medical Center, 3400 Spruce Street, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Alexander C
authors.initials: AC
authors.lastname: Wright
authors.affiliation: Sydney Microscopy &Microanalysis, University of Sydney, Sydney, New South Wales 2050, Australia.
authors.forename: Matthew
authors.initials: M
authors.lastname: Foley
authors.affiliation: Division of Cell Signaling and Immunology, University of Dundee, Dundee DD1 5EH, UK.
authors.forename: J Simon C
authors.initials: JS
authors.lastname: Arthur
authors.affiliation: Division of Cardiovascular Medicine and the Program in Molecular Medicine, University of Utah, Salt Lake City, Utah 84112, USA.
authors.forename: Kevin J
authors.initials: KJ
authors.lastname: Whitehead
authors.affiliation: Neurovascular Surgery Program, Section of Neurosurgery, Department of Surgery, The University of Chicago Medicine and Biological Sciences, Chicago, Illinois 60637, USA.
authors.forename: Issam A
authors.initials: IA
authors.lastname: Awad
authors.affiliation: Division of Cardiovascular Medicine and the Program in Molecular Medicine, University of Utah, Salt Lake City, Utah 84112, USA.|The Key Laboratory for Human Disease Gene Study of Sichuan Province, Institute of Laboratory Medicine, Sichuan Academy of Medical Sciences &Sichuan Provincial People's Hospital, Chengdu, Sichuan 610072, China.
authors.forename: Dean Y
authors.initials: DY
authors.lastname: Li
authors.affiliation: Laboratory of Cardiovascular Signaling, Centenary Institute, Sydney, New South Wales 2050, Australia.|Faculty of Medicine, Sydney Medical School, University of Sydney, Sydney, New South Wales 2050, Australia.
authors.forename: Xiangjian
authors.initials: X
authors.lastname: Zheng
authors.affiliation: Department of Medicine and Cardiovascular Institute, University of Pennsylvania, 3400 Civic Center Blvd, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Mark L
authors.initials: ML
authors.lastname: Kahn
mesh_terms: D051722:ADAM Proteins / Q000378:metabolism; D000818:Animals; D000831:Animals, Newborn; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D004195:Disease Models, Animal; D042783:Endothelial Cells / Q000201:enzymology / Q000378:metabolism*; D005260:Female; D020786:Hemangioma, Cavernous, Central Nervous System / Q000209:etiology / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors / Q000172:deficiency / Q000378:metabolism*; D048748:MAP Kinase Kinase Kinase 3 / Q000172:deficiency / Q000378:metabolism*; D020935:MAP Kinase Signaling System; D008297:Male; D051379:Mice; D011485:Protein Binding; D020741:rho GTP-Binding Proteins / Q000378:metabolism
chemical_list: C502544:CCM2 protein, human; D002352:Carrier Proteins; C096161:KLF2 protein, human; C000715467:KLF4 protein, human; C498314:Klf2 protein, mouse; C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; D048748:MAP Kinase Kinase Kinase 3; C482625:MAP3K3 protein, human; C482626:Map3k3 protein, mouse; D051722:ADAM Proteins; D020741:rho GTP-Binding Proteins