Two binding partners cooperate to activate the molecular motor Kinesin-1

The regulation of molecular motors is an important cellular problem, as motility in the absence of cargo results in futile adenosine triphosphate hydrolysis. When not transporting cargo, the microtubule (MT)-based motor Kinesin-1 is kept inactive as a result of a folded conformation that allows autoinhibition of the N-terminal motor by the C-terminal tail. The simplest model of Kinesin-1 activation posits that cargo binding to nonmotor regions relieves autoinhibition. In this study, we show that binding of the c-Jun N-terminal kinase–interacting protein 1 (JIP1) cargo protein is not sufficient to activate Kinesin-1. Because two regions of the Kinesin-1 tail are required for autoinhibition, we searched for a second molecule that contributes to activation of the motor. We identified fasciculation and elongation protein ζ1 (FEZ1) as a binding partner of kinesin heavy chain. We show that binding of JIP1 and FEZ1 to Kinesin-1 is sufficient to activate the motor for MT binding and motility. These results provide the first demonstration of the activation of a MT-based motor by cellular binding partners

Bayesian Modeling of the Yeast SH3 Domain Interactome Predicts Spatiotemporal Dynamics of Endocytosis Proteins

A genome-scale specificity and interaction map for yeast SH3 domain-containing proteins reveal how family members show selective binding to target proteins and predicts the dynamic localization of new candidate endocytosis proteins

An Elmo-Dock complex locally controls Rho GTPases and actin remodeling during cadherin-mediated adhesion

International audienceCell-cell contact formation is a dynamic process requiring the coordination of cadherin-based cell-cell adhesion and integrin-based cell migration. A genome-wide RNA interference screen for proteins required specifically for cadherin-dependent cell-cell adhesion identified an Elmo-Dock complex. This was unexpected as Elmo-Dock complexes act downstream of integrin signaling as Rac guanine-nucleotide exchange factors. In this paper, we show that Elmo2 recruits Dock1 to initial cell-cell contacts in Madin-Darby canine kidney cells. At cell-cell contacts, both Elmo2 and Dock1 are essential for the rapid recruitment and spreading of E-cadherin, actin reorganization, localized Rac and Rho GTPase activities, and the development of strong cell-cell adhesion. Upon completion of cell-cell adhesion, Elmo2 and Dock1 no longer localize to cell-cell contacts and are not required subsequently for the maintenance of cell-cell adhesion. These studies show that Elmo-Dock complexes are involved in both integrin- and cadherin-based adhesions, which may help to coordinate the transition of cells from migration to strong cell-cell adhesion

A genome-wide screen identifies conserved protein hubs required for cadherin-mediated cell-cell adhesion

International audienceCadherins and associated catenins provide an important structural interface between neighboring cells, the actin cytoskeleton, and intracellular signaling pathways in a variety of cell types throughout the Metazoa. However, the full inventory of the proteins and pathways required for cadherin-mediated adhesion has not been established. To this end, we completed a genome-wide (~14,000 genes) ribonucleic acid interference (RNAi) screen that targeted Ca(2+)-dependent adhesion in DE-cadherin-expressing Drosophila melanogaster S2 cells in suspension culture. This novel screen eliminated Ca(2+)-independent cell-cell adhesion, integrin-based adhesion, cell spreading, and cell migration. We identified 17 interconnected regulatory hubs, based on protein functions and protein-protein interactions that regulate the levels of the core cadherin-catenin complex and coordinate cadherin-mediated cell-cell adhesion. Representative proteins from these hubs were analyzed further in Drosophila oogenesis, using targeted germline RNAi, and adhesion was analyzed in Madin-Darby canine kidney mammalian epithelial cell-cell adhesion. These experiments reveal roles for a diversity of cellular pathways that are required for cadherin function in Metazoa, including cytoskeleton organization, cell-substrate interactions, and nuclear and cytoplasmic signaling

Early-Arriving Syp1p and Ede1p Function in Endocytic Site Placement and Formation in Budding Yeast

Recent studies have revealed the detailed timing of protein recruitment to endocytic sites in budding yeast. However, little is understood about the early stages of their formation. Here we identify the septin-associated protein Syp1p as a component of the machinery that drives clathrin-mediated endocytosis in budding yeast. Syp1p arrives at endocytic sites early in their formation and shares unique dynamics with the EH-domain protein Ede1p. We find that Syp1p is related in amino acid sequence to several mammalian proteins one of which, SGIP1-α, is an endocytic component that binds the Ede1p homolog Eps15. Like Syp1p, SGIP1-α arrives early at sites of clathrin-mediated endocytosis, suggesting that Syp1p/Ede1p and SGIP1-α/Eps15 may have a conserved function. In yeast, both Syp1p and Ede1p play important roles in the rate of endocytic site turnover. Additionally, Ede1p is important for endocytic site formation, whereas Syp1p acts as a polarized factor that recruits both Ede1p and endocytic sites to the necks of emerging buds. Thus Ede1p and Syp1p are conserved, early-arriving endocytic proteins with roles in the formation and placement of endocytic sites, respectively