We are using cell biological approaches to understand signaling mechanisms that govern the establishment and maintenance of epithelial cell polarity and their relevance for morphogenesis and cell transformation.
1. Identification of Par1b signaling pathways in epithelial cells - The role of Par1b in H. pylori pathology
Par1 isoforms are serine/threonine kinases that have emerged as "core determinants" of cell polarity in different contexts, including in mammalian epithelial cells. In collaboration with M. Stein’s group we found that Par1b is inactivated by a cytotoxic protein of Helicobacter pylori, a bacterium that destroys the epithelial lining of the stomach, resulting in gastritis and gastric ulcers and that is implicated in gastric cancer.
The relevant PAR1b- substrates and signaling pathways, however, are still poorly understood. We have conducted an unbiased screen that identified more than 70 putative novel Par1b substrates and their Par1b phosphorylation sites in polarized epithelial cells. We are employing a multi-tiered approach to elucidate their roles in epithelial morphogenesis and their relevance for H. pylori pathology utilizing immortalized epithelial cell lines and primary human gastric cell cultures.
2. Mechanisms for the establishment of hepatic versus columnar epithelial polarity
During development, simple (non-stratified) epithelia differentiate along two major lines: they either form lumina at their apices (e.g., columnar epithelia, such as kidney or intestine) or between the lateral membranes of neighboring cells (e.g., liver bile canaliculi -BC). Columnar and hepatic cells also employ different strategies for the biosynthetic protein delivery to the luminal surface. Columnar cells utilize a vectorial (direct) route from the transGolgi network (TGN) to the luminal membrane. On the other hand, liver cells utilize an indirect or transcytotic route, in which luminal markers are first targeted from the TGN to the basolateral surface, endocytosed, and further transported to the luminal domain. We have identified Par1b as the first candidate protein to contribute to this key branching of the epithelial differentiation program. Our goal is to gain insight into the underlying signaling mechanisms for distinct lumen position and trafficking phenotypes in columnar and hepatic cells by characterizing identified Par1 substrates.
Our repertoire of approaches include protein trafficking analysis in intact cells by time lapse imaging of fluorescently-tagged proteins, immunofluorescence and confocal laser microscopy to study cell morphology and sub-cellular protein distribution, biochemical assays that measure the kinetics of different protein trafficking steps and biochemical assays that measure protein-protein interactions and enzyme activities in cell lysates.
Cohen, D., Rodriguez-Boulan, E. and Müsch, A. (2004). "Par-1 promotes a hepatic mode of apical protein trafficking in MDCK cells". PNAS 101, 13792-97.
Cohen, D., Brennwald, P.J., Rodriguez-Boulan, E. and Müsch, A. (2004)."Mammalian PAR-1 determines epithelial lumen polarity by organizing the microtubule cytoskeleton". Journal Cell Biology, 164, 717-727.
Rodriguez-Boulan, E., Kreitzer G and Müsch, A. (2005) "Protein Sorting and Post-Golgi Trafficking in Epithelia."Nature Reviews Molecular Cell Biology, 6, 233-47.
Elbert, M., Cohen D.and Müsch, A, (2006): " Par1b promotes cell-cell adhesion and inhibits Dishevelled-mediated transformation of MDCK cells", Mol.Biol.Cell, 17, 3345-55.
Cohen D., Tian Y and Müsch, A (2007): "Par1b promotes hepatic-type lumen polarity in MDCK cells via myosin II and E-cadherin dependent signaling", Mol. Biol. Cell 18, 2203-15.
Zaher Zeaiter, David Cohen, Anne Müsch, Fabio Bagnoli, Antonello Covacci, and Markus Stein (2008) "Analysis of detergent resistant membranes of Helicobacter pylori infected gastric adenocarcinoma cells reveals a role for MARK2/Par1b in CagA-mediated disruption of cellular polarity", J Cellular Microbiology, 10, 781-94.
Cohen D., Fernandez D., Lazaro-Dieguez, F. and Anne Müsch,. “The serine/threonine kinase Par1b regulates epithelial lumen polarity via IRSp53-mediated cell-ECM signaling”, (2011) J Cell Biol. 192, 525-540.
Treyer A., Müsch, A. “Hepatiocyte Polarity” , (2013) Comprehensive Physiology, 3, 243-87.
Ispolatov I, Müsch, A. " A model for the self-organization of vesicular flux and protein distributions in the Golgi apparatus", (2013), PLOS Computational Biology, Jul;9(7):e1003125. doi: 10.1371/journal.pcbi.1003125
Lazaro-Dieguez, F. Cohen D., Fernandez D., Louis Hodgson, S. van IJzendoorn and Müsch, A. “Par1b links lumen polarity with LGN/NuMA positioning for distinct epithelial cell division phenotypes”, (2013) J Cell Biol. 203(2): 251-64
Slim CL, Lázaro-Diéguez F, Bijlard M, Toussaint MJ, de Bruin A, Du Q, Müsch A, van Ijzendoorn SC. Par1b induces asymmetric inheritance of plasma membrane domains via LGN-dependent mitotic spindle orientation in proliferating hepatocytes. PLoS Biol. 2013 Dec;11(12):e1001739.
Nachbar J, Lázaro-Diéguez F, Prekeris R, Cohen D, Müsch A. KIFC3 promotes mitotic progression and integrity of the central spindle in cytokinesis. Cell Cycle. 2014 13(3): 426-33
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Albert Einstein College of Medicine
Jack and Pearl Resnick Campus
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