How epithelial tissues make single-hole tubes

From a functional screen for genes required for 3D epithelial architecture we identify key roles for Synaptotagmin-like proteins 2 and 4 (Slp2-a/4-a) in generation of epithelial tubes with a single lumen. Together, Slp2-a/4-a co-ordinate the spatiotemporal organization of vectorial apical transport to ensure only a single apical surface, and thus formation of a single lumen, occurs per cell.

HFSP Career Development Award holder Fernando Martin-Belmonte and colleagues
authored on Tue, 17 July 2012

Epithelia are sheets of cells lining the body, both internally and externally, acting as selective barriers to separate our body from the outside. The epithelia were most possibly the first tissue that appeared during the evolution of multicellular animals, and is the first tissue that forms during embryogenesis. Epithelia constitute 60% of cells that form the human body, but are also the tissues where most of the tumors with the worst prognosis originate. In fact, 90% of all cancers  are derived from epithelial tissues (known as carcinomas). Epithelial cells are characterized by a high degree of cell polarity, with an apical cell membrane that separates the epithelium from the outside, and a basement membrane that connects these cells with blood vessels and the underlying extracellular matrix. One of the most important highlights in tumor progression is the loss of cell polarity.The formation of epithelial tissues requires both the generation of apical-basal polarity and the co-ordination of this polarity between neighboring cells to form a central lumen. During de novo lumen formation, vectorial membrane transport contributes to formation of a singular apical membrane, resulting in contribution of each cell to only a single lumen.

Figure: Morphogenesis of the epithelal lumen: different stages of MDCK cyst formation. A screening has identified Synaptotagmin-like protein 2-a (green) as a factor needed for coordination of apical membrane trafficking during lumen formation. Cysts stained with F-actin (red) and nuclei (blue).

The aim of this study was to identify new markers of epithelial tissue formation by a systematic screen with a genome-wide analysis and gene silencing using a 3D organotypic cell model, which is able to recapitulate the formation of epithelia "in vitro". This has served to characterize the molecular mechanisms by which epithelial tissues acquire cell polarity, or their "functional identity", during embryonic development,  and in diseases associated with this loss of polarity such as cancer .

The results of this research, to be published in Nature CellBiology in August 2012 , have identified 14 new essential components of the machinery for epithelial morphogenesis. Moreover, many of these markers are dramatically suppressed in tissue samples of epithelial tumors, indicating their potential importance as tumor markers. Then, we identify key roles for Synaptotagmin-like proteins 2-a and 4-a (Slp2-a/4-a) in generation of a single apical surface per cell. Slp2-a localizes to the luminal membrane in a PI(4,5)P2-dependent manner, where it targets vesicles to initiate a single lumen. On the other hand, vesicle tethering and fusion is controlled by Slp4-a, in conjunction with Rabs and SNARE proteins. In summary, Slp2-a/4-a co-ordinate the spatiotemporal organization of vectorial apical transport to ensure only a single apical surface, and thus formation of a single lumen, occurs per cell.

This work has been performed in a joint collaboration between Centro de Biología Molecular Severo Ochoa, a joint  Institute of the National Research Council (CSIC) and Universidad Autonoma de Madrid (Spain), with the University of California (USA) and the University Tohoku, Miyagi (Japan).



Figure: MDCK cyst phenotypes in an RNAi screening for lumen formation defects. Cysts stained with basolateral marker (green) and apical marker (red).


Synaptotagmin-Like Proteins Control Formation of a Single Apical Membrane Domain in Epithelial Cells.  Manuel Gálvez-Santisteban, Alejo E. Rodriguez-Fraticelli, David Bryant, Silvia Vergarajauregui, Takao Yasuda, Inmaculada Bañón-Rodríguez, Ilenia Bernascone, Anirban Datta, Natalie Spivak, Kitty Young, Christian Slim, Paul Breakman, Mitsunori Fukuda, Keith E. Mostov, and Fernando Martin-Belmonte. (2012) Nature Cell Biology doi:10.1038/ncb254.