Defining how Abelson Kinase Regulates Cell Adhesion and Actin Dynamics

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PROJECT SUMMARY The ability of cells to self-assemble into organs is extraordinary and requires tight coordination between the cell machinery modulating adhesion and the cytoskeleton. Defects in this coordination contribute to diseases ranging from developmental defects to cancer; thus it is critical to understand the mechanisms by which coordination of cell adhesion and the actin cytoskeleton is achieved. One regulator of this process is Abelson tyrosine kinase (Abl), which links receptors, small adaptor proteins, and cytoskeletal regulators. Abl kinases are key oncogenes and developmental regulators. Three models for Abl function currently exist: 1) Abl phosphorylates protein targets, thus altering their function and influencing cell behavior, 2) Abl directly modulates cytoskeletal dynamics through its conserved C-terminal actin binding domain (FABD), and 3) Abl acts as a scaffold to assemble multi-protein signaling complexes. However, the importance of each role during morphogenesis remains untested. To distinguish between these models, we tested mutant Abl proteins lacking kinase activity, F-actin binding, or motifs involved in protein interactions. Our preliminary data reveal that different mutants differentially affect distinct morphogenetic events from dorsal closure to apical constriction to CNS axon outgrowth. Strikingly, while kinase activity is required for full Abl activity, mutants lacking kinase activity or the FABD retain significant function during morphogenesis. Instead, a conserved PXXP motif within the linker region is more essential for Abl function during some aspects of morphogenesis than both kinase activity and the FABD. We also have identified candidate partners for the PXXP motif and found they affect processes similar to those affected by Abl. Thus, we hypothesize Abl functions as a robust regulatory machine during morphogenesis, with different aspects of its mechanisms of action (e.g., kinase activity, FABD domain, PXXP binding partners, etc.) being differentially important for certain biological processes. We will test this via the following specific aims: (1) Determine how specific functional domains of Abl differentially contribute to its function during morphogenesis using Drosophila dorsal closure (DC) as a model; (2) Determine how PXXP- binding partners work with Abl to regulate dynamic cell behaviors during DC. Specifically, we will use a combination of biochemical, genetic, and quantitative cell biological approaches to define how Abl's different mechanisms of action contribute to its ability to shape cell dynamics, organize functional linkages between the actin cytoskeleton and cell adhesions, and to shape regulatory interactions between multiple downstream actin regulators, including Ena, Dia, and Abi. In parallel, we will determine how Abl and its PXXP-binding partners (Abi or Crk) regulate dynamic cell behaviors during embryonic morphogenesis. This study will reveal how Abl and its interacting partners regulate cell adhesion and actin dynamics in a developmental context, providing new clues as to how Abl works to regulate dynamic cell behaviors during human development and disease.