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      A reduction of primary cilia but not hedgehog signaling disrupts morphogenesis in testicular organoids

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          Abstract

          Most mammalian cells possess a single, non-motile primary cilium that plays an important role in mediating cellular signaling pathways, such as Hedgehog (Hh) signaling. Primary cilia are present on testicular somatic cells and demonstrate a temporal expression during development; however, their role in testicular morphogenesis is not well characterized. To investigate the role of primary cilia and Hh signaling in Sertoli cells on morphogenesis, we inhibited assembly of primary cilia through CRISPR Cas9–mediated gene editing of ODF2, a structural component of primary cilia and siRNA-mediated gene silencing of IFT88, a functional component of the intraflagellar transport system. Knockdown of ODF2 and IFT88 resulted in a 50% reduction in the number of cells with primary cilia and significant shortening of the remaining cilia. The expression of GLI1, a downstream target of Hh signaling, was significantly reduced when IFT88 but not ODF2, was downregulated. When morphogenesis was examined using tubule formation in vitro and a novel testicular organoid system, loss of cilia after knockdown of both targets affected cellular assembly and organization. While the Hh pathway was found to be active during morphogenesis in vitro, addition of the Hh antagonist cyclopamine did not affect morphogenesis in either in vitro system. These results indicate that primary cilia are important for morphogenesis in vitro but Hh signaling is not the cilia-mediated pathway responsible for orchestrating morphogenic organization.

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          Author and article information

          Journal
          0417625
          2839
          Cell Tissue Res
          Cell Tissue Res
          Cell and tissue research
          0302-766X
          1432-0878
          12 January 2021
          03 January 2020
          April 2020
          19 January 2021
          : 380
          : 1
          : 191-200
          Affiliations
          [1 ]Faculty of Veterinary Medicine Comparative Biology and Experimental Medicine, University of Calgary, Calgary, Alberta, Canada
          [2 ]Cumming School of Medicine, Biochemistry and Molecular Biology, University of Calgary, 3330 Hospital Drive NW, Calgary, Alberta T2N4N1, Canada
          [3 ]Recombinetics Inc., 1246 University Avenue West, St. Paul, MN 55104, USA
          Author notes
          Article
          PMC7815324 PMC7815324 7815324 nihpa1659864
          10.1007/s00441-019-03121-8
          7815324
          31900662
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          Article

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