The long-term goal of this project is to understand the spatial and temporal regulation of signal transduction pathways that underlie mammalian cell polarization. Polarity establishment is vital for every aspect of metazoan development. For example, the segregation of cell fate determinants to the opposite poles of a stem cell, coupled with oriented cell division, permits the specification of distinct daughter cell phenotypes. Cell polarization is also required for morphogenesis, directional motility, antigen presentation, and axon guidance; and the loss of cell polarity is a critical step in cancer progression. Remarkably, the novel signaling pathways involved in polarization have been highly conserved throughout metazoan evolution. Cdc42 binds to a protein called Par6, and Par6 binds to atypical protein kinases C (aPKC). These proteins form a complex that regulates many types of cell polarization. We propose that Par6 behaves as a targeting subunit for aPKC, recruiting substrates for phosphorylation. Three putative effectors for Par6 have been found: Par3, Lgl, and Pals1. Each of these 3 proteins is essential for epithelial cell polarization, and each interacts with several other proteins that have also been implicated in polarization. These three sets of proteins (Par, Pals, and Lgl) interact both genetically and physically to create the distinct membrane domains that define a polarized epithelial cell, and they also participate in other types of cell polarity. How do they accomplish these tasks? The central questions to be addressed are: how do these proteins associate with and regulate each other? How do they localize to their appropriate positions during polarization (what are their targeting cues)? Which proteins need to arrive first at the cell junctions? And how do they execute the polarization program? X-ray crystallography will be used to determine the structures of polarity protein complexes. For cell biological assays, MDCK cells will be used as an established, well-accepted model system. These cells form highly polarized confluent monolayers, but in response to scatter factor, or wounding, they lose their apical/basal polarity, and become motile, with an anterior/posterior polarity axis Stable, inducible cell lines will be developed that express fluorescent fusions of polarity proteins, and used to quantify the dynamics of cell junction assembly and disassembly.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Cell Development and Function Integrated Review Group (CDF)
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Deatherage, James F
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University of Virginia
Schools of Medicine
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Ahmed, Syed Mukhtar; Nishida-Fukuda, Hisayo; Li, Yuchong et al. (2018) Exocyst dynamics during vesicle tethering and fusion. Nat Commun 9:5140
Ahmed, Syed Mukhtar; Macara, Ian G (2017) The Par3 polarity protein is an exocyst receptor essential for mammary cell survival. Nat Commun 8:14867
Ahmed, Syed Mukhtar; Macara, Ian G (2016) Mechanisms of polarity protein expression control. Curr Opin Cell Biol 42:38-45
Guyer, Richard A; Macara, Ian G (2015) Loss of the polarity protein PAR3 activates STAT3 signaling via an atypical protein kinase C (aPKC)/NF-?B/interleukin-6 (IL-6) axis in mouse mammary cells. J Biol Chem 290:8457-68
Iioka, Hidekazu; Macara, Ian G (2015) Detection of RNA-Protein Interactions Using Tethered RNA Affinity Capture. Methods Mol Biol 1316:67-73
Macara, Ian G; Guyer, Richard; Richardson, Graham et al. (2014) Epithelial homeostasis. Curr Biol 24:R815-25
Huo, Yongliang; Macara, Ian G (2014) The Par3-like polarity protein Par3L is essential for mammary stem cell maintenance. Nat Cell Biol 16:529-37
Rodriguez-Boulan, Enrique; Macara, Ian G (2014) Organization and execution of the epithelial polarity programme. Nat Rev Mol Cell Biol 15:225-42
Andreeva, Anna; Lee, Jianyi; Lohia, Madhura et al. (2014) PTK7-Src signaling at epithelial cell contacts mediates spatial organization of actomyosin and planar cell polarity. Dev Cell 29:20-33
Baschieri, Francesco; Confalonieri, Stefano; Bertalot, Giovanni et al. (2014) Spatial control of Cdc42 signalling by a GM130-RasGRF complex regulates polarity and tumorigenesis. Nat Commun 5:4839

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