Uncovering the Role of Protein Kinase A in Dictyostelium Chemotaxis toward Cyclic Adenosine Monophosphate

Abstract

Chemotaxis, or directed cell migration, is important in many biological processes such as embryonic development, wound healing, and the direction of immune cells to sites of inflammation or infection. When not regulated properly, chemotaxis is implicated in disease states including inflammatory diseases and cancer metastasis. During eukaryotic chemotaxis, cells are able to sense a chemical gradient through receptors on the cell membrane that trigger complicated intracellular signaling networks, ultimately resulting in changes in the actin cytoskeleton leading to cell migration. The proteins involved in these signaling networks require tight spatiotemporal regulation, and the mechanisms underlying this regulation are not well understood. The work of this dissertation aims to better elucidate the pathways that regulate chemotaxis and enable cells to respond and adapt to changes in the chemoattractant gradient. To this end, we utilized the model organism Dictyostelium discoideum, and focused on determining the role of protein kinase A (PKA) in regulating the chemotactic pathways. We found that, in cells lacking PKA activity, key proteins upstream of PKA are deregulated, which suggests that PKA acts in one or more negative feedback loops to inactivate the chemotactic pathways. To narrow in on potential direct sites of PKA regulation, we studied the effects of lack of PKA activity on G-protein dissociation, and we also analyzed the chemotactic role of potential PKA phosphorylation sites of a Ras family protein. In addition to these studies, we also began an investigation into the similarities and potential cross-talk between PKA and a relatively novel chemotactic player, Myosin G. Overall, we have identified new roles for PKA in Dictyostelium chemotaxis, and have identified potential mechanisms in which PKA negatively regulates the chemotactic pathways, which would aid in the cells ability to efficiently respond to changes in its surrounding signal gradient.