Characterizing membrane receptor signaling: New roles for electrostatic residues in EGFR regulation and actin in FcεRI signaling.
Mohr, Jordan David
Membrane proteins play a critical role in cellular survival, function, and communication. Membrane receptors are one of the primary means by which cells interact with their extracellular environment to locate nutrients, identify threats, and communicate with other cells. This communication across the cell membrane is referred to as signal transduction and represents an immense biophysical feat that has evolved numerous transduction mechanisms. Given the importance of signal transduction, it is no surprise that defects in membrane receptor function have been linked to many disease states in humans. The membrane receptors discussed in this work are the epidermal growth factor receptor (EGFR) and the immunoglobulin E (IgE) FcεRI immunorececptor. Dysregulation of EGFR through gain-of-function mutations has been linked to many cancers and is most commonly observed in brain, breast, and non-small cell lung cancer, while activation of FcεRI through normally benign antigens triggers the allergic response. Both of these represent cases of inappropriate receptor activation, though through differing mechanisms. Both of these receptors also serve as model receptors with broad implications to other receptors: EGFR is a model receptor tyrosine kinase while FcεRI is a model immunoreceptor that shares many similarities with B-cell and T-cell receptor signaling. Improving our understanding of these receptors also benefits our understanding of these other systems and the pathologies associated with their dysregulation. An earlier study of ours demonstrated that electrostatic residues in the juxtamembrane domain of EGFR play an inhibitory role by preventing spontaneous receptor activation. Prompted by this, we attempted to identify a potential mechanism for this activation. We were able to show that this mutation requires a series of conformational states consistent with an “inside-out” activation mechanism, in which these electrostatic residues prevent receptor dimerization and activation in the absence of ligand. In another study, we showed that we could use microfabricated patterned ligand features to study protein recruitment to FcεRI clusters and quantify the effects of actin polymerization inhibitors on this recruitment. Here we expand upon that work by showing the recruitment of all the components of the Arp2/3 signaling complex in RBL-2H3 mast cells on patterned features. We also identify new roles for Arp2/3 and formins in FcεRI activation using pharmacological inhibitors. These actin-catalyzing enzymes appear to play both cooperative and differential roles in FcεRI signaling, with Arp2/3 potentially mediating complex stability while formins maintain quality control of the basal actin cytoskeleton and cellular processes. This is the first time that these roles have been investigated in detail in FcεRI signaling. Taken together, this work extends our knowledge of juxtamembrane electrostatic residues in EGFR regulation and identifies new, differential roles for actin-polymerizing enzymes in FcεRI signaling.
Actin; EGFR; Electrostatic; FcεRI; Microfabrication
Baird, Barbara A.
Holowka, David Allen; Lin, Hening; Cerione, Richard A.
Biomedical and Biological Sciences
Ph. D., Biomedical and Biological Sciences
Doctor of Philosophy
Attribution-NonCommercial-ShareAlike 4.0 International
dissertation or thesis
Except where otherwise noted, this item's license is described as Attribution-NonCommercial-ShareAlike 4.0 International