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Published 6 August 2001. doi:10.1083/jcb.200104057
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© The Rockefeller University Press, 0021-9525/2001/8/549 $5.00
The Journal of Cell Biology, Volume 154, Number 3, August 6, 2001 549-576

Article

A protein interaction map for cell polarity development



Becky L. Drees2, Bryan Sundin4, Elizabeth Brazeau2, Juliane P. Caviston5, Guang-Chao Chen8, Wei Guo12, Keith G. Kozminski1, Michelle W. Lau1, John J. Moskow9, Amy Tong6, Laura R. Schenkman7, Amos McKenzie, III7, Patrick Brennwald10, Mark Longtine11, Erfei Bi5, Clarence Chan8, Peter Novick13, Charles Boone6, John R. Pringle7, Trisha N. Davis4, Stanley Fields3 and David G. Drubin1

1 Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720
2 Departments of Genetics and Medicine, University of Washington, Seattle, WA 98195
3 Howard Hughes Medical Institute and Departments of Genetics and Medicine, University of Washington, Seattle, WA 98195
4 Department of Biochemistry, University of Washington, Seattle, WA 98195
5 Department of Cell and Developmental Biology, University of Pennsylvania School of Medicine, Philadelphia, PA 19104
6 Banting and Best Department of Medical Research, University of Toronto, Toronto, Ontario M5G 1L6, Canada
7 Department of Biology, University of North Carolina, Chapel Hill, NC 27599
8 Section of Molecular Genetics and Microbiology, Institute for Cellular and Molecular Biology, University of Texas, Austin, TX 78712
9 Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, NC 27710
10 Department of Cell Biology and Genetics, Cornell University, New York, NY 10021
11 Department of Biochemistry and Molecular Biology, Oklahoma State University, Stillwater, OK 74078
12 Biology Department, University of Pennsylvania, Philadelphia, PA 19104
13 Department of Cell Biology, Yale University School of Medicine, New Haven, CT 06520

Address correspondence to David Drubin, Department of Molecular and Cell Biology, 401 Barker Hall, University of California, Berkeley, CA 94720-3202. Tel.: (510) 642-3692. Fax: (510) 643-0062. E-mail: drubin{at}uclink4.berkeley.edu

Many genes required for cell polarity development in budding yeast have been identified and arranged into a functional hierarchy. Core elements of the hierarchy are widely conserved, underlying cell polarity development in diverse eukaryotes. To enumerate more fully the protein–protein interactions that mediate cell polarity development, and to uncover novel mechanisms that coordinate the numerous events involved, we carried out a large-scale two-hybrid experiment. 68 Gal4 DNA binding domain fusions of yeast proteins associated with the actin cytoskeleton, septins, the secretory apparatus, and Rho-type GTPases were used to screen an array of yeast transformants that express ~90% of the predicted Saccharomyces cerevisiae open reading frames as Gal4 activation domain fusions. 191 protein–protein interactions were detected, of which 128 had not been described previously. 44 interactions implicated 20 previously uncharacterized proteins in cell polarity development. Further insights into possible roles of 13 of these proteins were revealed by their multiple two-hybrid interactions and by subcellular localization. Included in the interaction network were associations of Cdc42 and Rho1 pathways with proteins involved in exocytosis, septin organization, actin assembly, microtubule organization, autophagy, cytokinesis, and cell wall synthesis. Other interactions suggested direct connections between Rho1- and Cdc42-regulated pathways; the secretory apparatus and regulators of polarity establishment; actin assembly and the morphogenesis checkpoint; and the exocytic and endocytic machinery. In total, a network of interactions that provide an integrated response of signaling proteins, the cytoskeleton, and organelles to the spatial cues that direct polarity development was revealed.

Key Words: cytoskeleton; Rho proteins; secretion; cell polarity; endocytosis


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