TGF beta regulation of cilium-dependent signaling

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DESCRIPTION (provided by applicant): The primary cilium plays important roles in cellular signaling and mammalian development. Cilia defects are associated with a number of human developmental disorders, including hydrocephaly, chronic pulmonary obstruction, polycystic kidney disease and left-right asymmetry defects. Primary cilia are critical for Sonic hedgehog (Shh) signaling, and may play roles in other signaling pathways. Mutations that disrupt Shh signaling or cilia function cause similar phenotypes in mice, including limb defects and holoprosencephaly (HPE), a severe genetic disorder affecting craniofacial development. Loss of function mutations in the human genes encoding Shh and Tgif1 are associated with HPE. Tgif1 is a transcriptional corepressor that limits transforming growth factor (TGF) signaling via Smad2. However, it is not known how Tgif1, and the related Tgif2, regulate embryogenesis, or how Tgifs and TGF signaling regulate ciliogenesis and Shh signaling. We propose to test the model that TGF family signaling must be strictly limited during embryonic development, and that unrestrained TGF family signaling via Smad2 disrupts cell polarity and the formation of primary cilia, thereby inhibiting cilia-dependent signaling. By conditionally targeting both the Tgif1 and Tgif2 genes in mice, we have created a mouse model for excess TGF signaling, in which cell polarity and ciliogenesis are disrupted. Additionally, mice lacking all Tgif function have embryonic defects, including HPE and left-right asymmetry defects, and have increased expression of the pluripotency factor, Nanog. Specifically, we will: 1) Determine whether Tgifs control Shh signaling in the forebrain by restraining TGF/Nodal signaling via Smad2, and test whether HPE in the absence of Tgifs is due to excess TGF family signaling. 2) Determine whether, in the absence of Tgif function, the forebrain is unable to respond to Shh signaling due to polarity defects and the absence of primary cilia, or whether Tgifs and TGF signaling directly regulate Shh gene expression. 3) Determine whether TGF family signaling must be limited to prevent disruption of primary cilia and cilia- dependent signaling in other embryonic tissues, including the neural tube and node. Finally, we will test whether Nanog is a direct target for repression by Tgifs, and whether excess Nanog expression causes the cell polarity and ciliogenesis defects. This work will determine how Tgifs and TGF signaling are integrated with the Shh pathway, and determine the role that TGF signaling plays in ciliogenesis and cilia-dependent signaling.