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First published online October 22, 2008
doi: 10.1242/10.1242/jcs.018648

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ROCK-mediated contractility, tight junctions and channels contribute to the conversion of a preapical patch into apical surface during isochoric lumen initiation

Aldo Ferrari^1,*,, Alexey Veligodskiy^1,2,, Ulrich Berge^1,2,, Miriam S. Lucas³ and Ruth Kroschewski^1,

¹ Institute of Biochemistry, ETH Zurich, Schafmattstrasse 18, 8093 Zurich, Switzerland
² Molecular Life Science Ph.D. Program, Zurich, Switzerland
³ Electron Microscopy ETH Zurich (EMEZ), Wolfgang-Pauli-Strasse 16, 8093 Zurich, Switzerland

Author for correspondence (e-mail: kroschewski{at}bc.biol.ethz.ch)

Accepted 31 August 2008

Epithelial cells assemble into three-dimensional aggregates to generate lumen-containing organ substructures. Cells therein contact the extracellular matrix with their basal surface, neighbouring cells with their contact surface and the lumen with their apical surface. We investigated the development of single MDCK cells into aggregates with lumen using quantitative live-cell imaging to identify morphogenetic rules for lumen formation. In two-cell aggregates, membrane insertion into the contact surface established a preapical patch (PAP) characterized by the presence of the apical marker gp135, microvilli and the absence of E-cadherin. This PAP originated from a compartment that had hallmarks of an apical recycling endosome, and matured through Brefeldin-A-sensitive membrane trafficking and the establishment of tight junctions around itself. As a result of the activity of water and ion channels, an optically resolvable lumen formed. Initially, this lumen enlarged without changes in aggregate volume or cell number but with decreasing cell volumes. Additionally, the ROCK1/2-myosin-II pathway counteracted PAP and lumen formation. Thus, lumen formation results from PAP establishment, PAP maturation, lumen initiation and lumen enlargement. These phases correlate with distinct cell surface and volume patterns, which suggests that such morphometric parameters are regulated by trafficking, ROCK-mediated contractility and hydrostatic pressure or vice versa.

Key words: Epithelial cell polarity, Cell shape, Topology, Surface surveillance, Morphogenesis, 3D cell culture, Volume, Force