title: Structure of a highly conserved domain of Rock1 required for Shroom-mediated regulation of cell morphology.
abstract: Rho-associated coiled coil containing protein kinase (Rho-kinase or Rock) is a well-defined determinant of actin organization and dynamics in most animal cells characterized to date. One of the primary effectors of Rock is non-muscle myosin II. Activation of Rock results in increased contractility of myosin II and subsequent changes in actin architecture and cell morphology. The regulation of Rock is thought to occur via autoinhibition of the kinase domain via intramolecular interactions between the N-terminus and the C-terminus of the kinase. This autoinhibited state can be relieved via proteolytic cleavage, binding of lipids to a Pleckstrin Homology domain near the C-terminus, or binding of GTP-bound RhoA to the central coiled-coil region of Rock. Recent work has identified the Shroom family of proteins as an additional regulator of Rock either at the level of cellular distribution or catalytic activity or both. The Shroom-Rock complex is conserved in most animals and is essential for the formation of the neural tube, eye, and gut in vertebrates. To address the mechanism by which Shroom and Rock interact, we have solved the structure of the coiled-coil region of Rock that binds to Shroom proteins. Consistent with other observations, the Shroom binding domain is a parallel coiled-coil dimer. Using biochemical approaches, we have identified a large patch of residues that contribute to Shrm binding. Their orientation suggests that there may be two independent Shrm binding sites on opposing faces of the coiled-coil region of Rock. Finally, we show that the binding surface is essential for Rock colocalization with Shroom and for Shroom-mediated changes in cell morphology.
authors.affiliation: Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
authors.forename: Swarna
authors.initials: S
authors.lastname: Mohan
authors.affiliation: Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
authors.forename: Debamitra
authors.initials: D
authors.lastname: Das
authors.affiliation: Department of Chemistry, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Bauer
authors.affiliation: Department of Biology, Brookhaven National Laboratory, Upton, New York, United States of America.
authors.forename: Annie
authors.initials: A
authors.lastname: Heroux
authors.affiliation: Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
authors.forename: Jenna K
authors.initials: JK
authors.lastname: Zalewski
authors.affiliation: Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
authors.forename: Simone
authors.initials: S
authors.lastname: Heber
authors.affiliation: Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
authors.forename: Atinuke M
authors.initials: AM
authors.lastname: Dosunmu-Ogunbi
authors.affiliation: Department of Chemistry, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Trakselis
authors.affiliation: Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
authors.forename: Jeffrey D
authors.initials: JD
authors.lastname: Hildebrand
authors.affiliation: Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
authors.forename: Andrew P
authors.initials: AP
authors.lastname: Vandemark
mesh_terms: D005454:Fluorescence Polarization; D005455:Fluorescent Antibody Technique; D006801:Humans; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism*; D024462:Myosin Type II / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D054460:rho-Associated Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D008840:Microfilament Proteins; C515851:ROCK1 protein, human; D054460:rho-Associated Kinases; D024462:Myosin Type II