IQGAP1 is a conserved modular protein overexpressed in cancer and involved in organizing actin and microtubules in motile processes such as adhesion, migration, and cytokinesis. A variety of proteins have been shown to interact with IQGAP1, including the small G proteins Rac1 and Cdc42, actin, calmodulin, ␤-catenin, the microtubule plus end-binding proteins CLIP170 (cytoplasmic linker protein) and adenomatous polyposis coli. However, the molecular mechanism by which IQGAP1 controls actin dynamics in cell motility is not understood. Quantitative co-localization analysis and down-regulation of IQGAP1 revealed that IQGAP1 controls the co-localization of N-WASP with the Arp2/3 complex in lamellipodia. Co-immunoprecipitation supports an in vivo link between IQGAP1 and N-WASP. Pull-down experiments and kinetic assays of branched actin polymerization with N-WASP and Arp2/3 complex demonstrated that the C-terminal half of IQGAP1 activates N-WASP by interacting with its BR-CRIB domain in a Cdc42-like manner, whereas the N-terminal half of IQGAP1 antagonizes this activation by association with a C-terminal region of IQGAP1. We propose that signal-induced relief of the autoinhibited fold of IQGAP1 allows activation of N-WASP to stimulate Arp2/3-dependent actin assembly.Directional cell migration results from the coordination of protrusion formation and cell adhesion. Although the concerted re-organization of actin and microtubules establishes and maintains cell polarization during directional movement, little is known about the molecular mechanisms underlying signal-mediated crosstalk between the two different cytoskeletal arrays (1). In this context, the modular IQGAP1 protein has received intense interest in the past years (2). The multiple partners of IQGAP1, including signaling molecules like Cdc42 or Rac1, calmodulin (3-6), and adhesion/cytoskeletal proteins like ␤-catenin, E-cadherin, actin filaments, and microtubule plus end-tracking proteins (CLIP170 and adenomatous polyposis coli (APC)) strongly suggest that IQGAP1 is an important player in coordinating cell polarity, adhesion, and migration (7-13). Concrete support to this view was brought by evidence showing that IQGAP1 is overexpressed in cancer (14, 15), controls cytokinesis (16 -21), and cell-cell adhesion (22-24). In addition, recent reports showed that IQGAP1 localizes in lamellipodia of motile cells (4,25,26) where it may link microtubule ends to the actin cytoskeleton (12,27) and that overexpression of IQGAP1 increases cell motility, whereas knockdown of the protein reduces cell migration and inhibits the formation of a protrusive actin meshwork at the leading edge (25). Finally, IQGAP1 regulates E-cadherinmediated cell-cell adhesion both positively and negatively (11). However, the functional and molecular link between IQGAP1 and the actin cytoskeleton in cell-cell adhesions and in lamellipodia has remained elusive.Extension of lamellipodia is driven by stimulus-responsive WASP family proteins (N-WASP, WASP, and Scar/WAVE) Cortactin and CARMIL, which act...

SummaryROCK-mediated contractility, tight junctions and channels contribute to the conversion of a preapical patch into apical surface during isochoric lumen initiation

Mechanical interactions between cells and extracellular matrix (ECM) mediate epithelial cyst formation. This work relies on the combination of numerical modeling with live cell imaging, to piece together a novel nonintrusive method for determining three-dimensional (3D) mechanical forces caused by shape changes of a multicellular aggregate at the early stages of epithelial cyst formation. We analyzed the evolution of Madin-Darby canine kidney cells in 3D cultures using time-lapse microscopy, with type I collagen gel forming the ECM. The evolving 3D interface between the ECM and the cell aggregate was obtained from microscopy images, and the stress on the surface of a proliferating aggregate and in the surrounding ECM was calculated using the finite element method. The viscoelastic properties of the ECM (a needed input for the finite element method solver) were obtained through oscillatory shear flow experiments on a rheometer. For validation purpose, the forces exerted by an aggregate on a force-sensor array were measured and compared against the computational results.