One mechanism by which branched epithelial structures can be generated involves the formation of a cleft that splits an epithelial sheet into separate buds. Wedge-shaped accumulations of the extracellular matrix (ECM) protein fibronectin appear at sites of cleft formation and persist at the base of the cleft as it progresses. Cells adjacent to the fibronectin accumulations lose the adhesion protein E-cadherin from their surfaces, leading to the hypothesis that clefts are created by transient formation of cell-matrix adhesions at the expense of cell-cell adhesions. Onodera et al. identified btbd7 (BTB domain 7) in a screen for genes transcriptionally up-regulated in clefts of actively branching embryonic murine salivary glands, where its mRNA was concentrated near sites of fibronectin accumulation. In salivary gland epithelial explants, purified fibronectin, but not other ECM components, induced btbd7 expression and disorganization of the outer layer of epithelial cells, a morphological change that was rescued by small interfering RNA (siRNA) depletion of Btbd7. In two- and three-dimensional cultures of the Madin-Darby canine kidney (MDCK) epithelial cell line, expression of transgenic btbd7 reduced E-cadherin abundance at cell-cell boundaries, increased cell dispersal and migration speed, and induced expression of Snail2, which encodes a transcription factor implicated in cell dispersal. Snail2 transcripts were enriched near clefts in vivo and up-regulated in salivary gland epithelia explants cultured with fibronectin. Treating explants with siRNAs targeting btbd7 prevented fibronectin-induced Snail2 up-regulation, but siRNAs targeting Snail2 did not affect btbd7 expression, indicating that the Btbd7 was upstream of Snail2. Treating cultured salivary glands with siRNAs targeting btbd7 or Snail2 reduced the frequency of cleft formation and reduced the speed of cleft progression, whereas overexpression of btbd7 in salivary gland explants increased spreading and cell dispersal. btbd7 was also expressed near sites of fibronectin accumulation in clefts of branching mouse embryonic lungs, and btbd7 siRNA reduced branching in cultured lungs. This sequence of events whereby fibronectin induces production of Btbd7, which, in turn, induces Snail2 expression and suppresses E-cadherin accumulation, is consistent with a model in which transient loss of cell-cell interactions enables cleft formation and propagation.
T. Onodera, T. Sakai, J. C.-f. Hsu, K. Matsumoto, J. A. Chiorini, K. M. Yamada, Btbd7 regulates epithelial cell dynamics and branching morphogenesis. Science 329, 562–565 (2010). [Abstract] [Full Text]