The regulation of cell architecture and morphology is essential during the course of vertebrate development, organogenesis, and tissue homeostasis. The actin-associated Shroom proteins are critical determinants of epithelial cell shape, controlling the process of apical constriction and cellular contractility. Shroom proteins work by targeting the Rock-Myosin II pathway to specific regions of the cells where it causes the formation of a contractile actomyosin network. The ability of Shroom to engage the Rock-myosin pathway is dependent on a conserved domain that binds directly to Rock. Based on our work, we predict that this pathway represents an evolutionarily conserved signaling module that is required for a number of developmental events in vertebrates, including formation of the central nervous system, eye, and intestines. This proposal will investigate the assembly and function of this pathway at the structural, biochemical, and cellular levels. We will accomplish this through the following aims.
Aim I. Structural Analysis and Characterization of a Shrm SD2 domain. We will use X-ray crystallography to determine the structure for an SD2 domain as a starting point for generating hypotheses about how SD2 interacts with Rock to control contractility. We will use biochemical and cell biological approaches, combined with mutants derived from our analysis of the structure, to test these hypotheses.
Aim II. Characterization of the Shroom-Binding Domain (SBD) of Rock. We will determine the structure of SBD to give insight into its conformation and design informative mutants to determine which portions of SBD are physically interacting with Shrm. We will also examine the effects of these mutations in vivo. Coupled with the structure of SD2, we will be able to generate simple testable models that describe the SD2-SBD interaction and how it regulates Rock activity.
Aim III. Structure of the SD2-SBD complex and implications for signaling. The interaction between Shrm and Rock is sufficient to regulate downstream cytoskeletal changes and alter cell morphology. We will address the nature of the interaction by solving the structure of the SD2- SBD complex. We will expand our analysis to examine the role each domain plays in regulating cell morphology and analyze the interplay between the Shrm-Rock pathway and the Rho-Rock pathway. These studies are significant because only by elucidating how signaling complexes are assembled can we understand their manner of function and regulation. Because the Rock- myosin II pathway is central to a vast number of cellular processes, understanding how this pathway functions may provide invaluable insight into the basic mechanisms of cellular regulation under both normal and disease conditions. In addition, targeted disruption of specific Rock-dependent events may prove to be a powerful therapeutic approach to treating certain human diseases and disorders.
Cells must be able to regulate their shape in order to form the correct body plan and functional tissues and organs. This research investigates a network of proteins that controls how cells manage this complicated task. By understanding these processes from the atomic to the cellular level we hope to learn how defects in these pathways may cause human disease.
|Zalewski, Jenna K; Heber, Simone; Mo, Joshua H et al. (2017) Combining Wet and Dry Lab Techniques to Guide the Crystallization of Large Coiled-coil Containing Proteins. J Vis Exp :|
|Wenzel, Sally E; Tyurina, Yulia Y; Zhao, Jinming et al. (2017) PEBP1 Wardens Ferroptosis by Enabling Lipoxygenase Generation of Lipid Death Signals. Cell 171:628-641.e26|
|Zalewski, Jenna K; Mo, Joshua H; Heber, Simone et al. (2016) Structure of the Shroom-Rho Kinase Complex Reveals a Binding Interface with Monomeric Shroom That Regulates Cell Morphology and Stimulates Kinase Activity. J Biol Chem 291:25364-25374|
|Roland, Bartholomew P; Zeccola, Alison M; Larsen, Samantha B et al. (2016) Structural and Genetic Studies Demonstrate Neurologic Dysfunction in Triosephosphate Isomerase Deficiency Is Associated with Impaired Synaptic Vesicle Dynamics. PLoS Genet 12:e1005941|
|Roland, Bartholomew P; Amrich, Christopher G; Kammerer, Charles J et al. (2015) Triosephosphate isomerase I170V alters catalytic site, enhances stability and induces pathology in a Drosophila model of TPI deficiency. Biochim Biophys Acta 1852:61-9|
|McGreevy, Erica M; Vijayraghavan, Deepthi; Davidson, Lance A et al. (2015) Shroom3 functions downstream of planar cell polarity to regulate myosin II distribution and cellular organization during neural tube closure. Biol Open 4:186-96|
|Das, Debamitra; Zalewski, Jenna K; Mohan, Swarna et al. (2014) The interaction between Shroom3 and Rho-kinase is required for neural tube morphogenesis in mice. Biol Open 3:850-60|
|Mohan, Swarna; Das, Debamitra; Bauer, Robert J et al. (2013) Structure of a highly conserved domain of Rock1 required for Shroom-mediated regulation of cell morphology. PLoS One 8:e81075|
|Roland, Bartholomew P; Stuchul, Kimberly A; Larsen, Samantha B et al. (2013) Evidence of a triosephosphate isomerase non-catalytic function crucial to behavior and longevity. J Cell Sci 126:3151-8|
|Celotto, Alicia M; Liu, Zhaohui; Vandemark, Andrew P et al. (2012) A novel Drosophila SOD2 mutant demonstrates a role for mitochondrial ROS in neurodevelopment and disease. Brain Behav 2:424-34|
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