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Dissection of the Role of CCM Genes in Tubulogenesis Using the Drosophila Tracheal System as a Model

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Cerebral Cavernous Malformations (CCM)

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2152))

Abstract

Embryos deficient for an essential gene may show complex phenotypes that reflect pleiotropic functions and non-cell-autonomous requirements for the encoded protein. The generation of mosaic animals, where most cells are wild type, but a few cells are mutant, is a powerful tool permitting the detailed analysis of the cell autonomous function of a gene, in a particular cell type, at cellular and subcellular resolutions. Here we apply this method to the analysis of the Cerebral Cavernous Malformations 3 (CCM3) pathway in Drosophila.

The conserved CCM3 protein functions together with its binding partner, Germinal Center Kinase III (Wheezy/GckIII in Drosophila, MST3, STK24, and STK25 in human) in the regulation of tube morphogenesis (Bergametti et al. Am J Hum Genet. 76:42–51, 2005; Fidalgo et al. J Cell Sci. 123:1274–1284, 2010; Guclu et al. Neurosurgery. 57:1008–1013, 2005; Lant et al. Nat Commun. 6:6449, 2015; Song et al. Dev Cell. 25:507–519, 2013; Ceccarelli et al. J Biol Chem. 286:25056–25064, 2011; Rehain-Bell et al. Curr Biol. 27:860–867, 2017; Xu et al. Structure. 21:1059–1066, 2013; Zhang et al. Front Biosci. 17:2295–2305, 2012; Zhang et al. Dev Cell. 27:215–226, 2013; Zheng et al. J Clin Invest. 120:2795–2804, 2010). The Drosophila proteins play a role in the regulation of tube shape in the tracheal (respiratory) system, analogous to the role of the human proteins in the vascular system. To understand the cellular basis for tube dilation defects caused by loss of pathway function, we describe techniques for the generation and analysis of positively marked homozygous mutant GckIII tracheal cells, coupled with an “open book” preparation that can be subjected to immunofluorescent analysis. Dozens of mutant tracheal cells are generated per mosaic animal, and neighboring heterozygous cells in the same animal serve as ideal internal controls.

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Authors and Affiliations

  1. The Taub Institute, New York, NY, USA

    Alondra B. Schweizer Burguete

  2. The Department of Pathology and Cell Biology, Columbia University Medical Center, New York, NY, USA

    Amin S. Ghabrial

Authors
  1. Alondra B. Schweizer Burguete
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  2. Amin S. Ghabrial
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Corresponding author

Correspondence to Amin S. Ghabrial .

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Editors and Affiliations

  1. Department of Biotechnology, Chemistry and Pharmacy, University of Siena, SIENA, Italy

    Lorenza Trabalzini

  2. Department of Biotechnology, Chemistry and Pharmacy, University of Siena, SIENA, Italy

    Federica Finetti

  3. Department of Clinical and Biological Sciences, School of Medicine and Surgery, University of Torino, ORBASSANO (TORINO), Italy

    Saverio Francesco Retta

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Schweizer Burguete, A.B., Ghabrial, A.S. (2020). Dissection of the Role of CCM Genes in Tubulogenesis Using the Drosophila Tracheal System as a Model. In: Trabalzini, L., Finetti, F., Retta, S. (eds) Cerebral Cavernous Malformations (CCM) . Methods in Molecular Biology, vol 2152. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0640-7_14

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  • DOI: https://doi.org/10.1007/978-1-0716-0640-7_14

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-0639-1

  • Online ISBN: 978-1-0716-0640-7

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