CHE Fall Seminar Series

Details

Speaker: Lawrence A. Stern
Assistant Professor
Department of Chemical, Biological and Materials Engineering
University of South Florida

Title: Engineering and Applying Intracellular Co-Localization to Study Cell Signaling

Abstract:
Cells sense cues in their environments and change their behavior to respond using cascades of protein-protein and protein-molecule interactions collectively known as cell signaling. Often, these signaling cascades are driven by post-translational modifications that are the result of co-localizing two proteins in a contingent manner. These responses play important roles in health and disease, making them prime targets for study and engineering, with a goal to modulate cell behavior for therapeutic benefit. However, major gaps remain in the techniques used to study and engineer intracellular co-localization and the subsequent post-translational modifications that follow. In this seminar, I will discuss my lab’s efforts to address these challenges through the development and application of new high-throughput screening platforms using yeast. First, I will describe our recent work in the characterization of kinase-substrate interaction resulting in substrate phosphorylation by adapting a previously described endoplasmic reticulum sequestration screening strategy with yeast surface display. These phosphorylation events are the currency of cell signaling, the understanding of which is imperative both for mapping natural cell signaling networks and for successful synthetic receptor engineering. We have demonstrated the ability to 1) display full-length receptor intracellular domains on the yeast surface, 2) robustly enrich phosphorylated intracellular domains from dilute pools using magnetic selection, 3) profile substrate specificities for a tyrosine kinase, and we have explored building 2-kinase phosphorylation cascades in the yeast ER. Recent progress in extending these techniques toward cell-surface substrate modification and engineering biased signaling modules will be discussed. Second, I will describe our efforts in developing a new co-localization platform in yeast that leverages naturally occurring membrane-associating domains to bring proteins of interest to the cytoplasmic side of the cell membrane. This compartment is where many protein-protein interactions that initiate cell signaling cascades occur and thus is the ideal environment to study these interactions. We use split fluorescent proteins to validate this co-localization approach, study different combinations of protein fusions to optimize co-localization, and extend this to demonstrate that the co-localization system can distinguish between “bright” and “dim” fluorescent protein phenotypes in a library setting.

Bio:
Dr. Lawrence A. Stern earned Bachelor of Science degrees in Chemical Engineering and Chemistry at Virginia Tech and a Ph.D. in Chemical Engineering at the University of Minnesota under the mentorship of Dr. Ben Hackel. He completed postdoctoral study in the T Cell Therapeutics Research Laboratory at City of Hope under the mentorship of Dr. Christine Brown and Dr. Stephen Forman. After completing his studies, Dr. Stern began his independent research career in January 2020 at the University of South Florida as an Assistant Professor in the Department of Chemical, Biological and Materials Engineering. His lab applies protein engineering and high-throughput screening techniques to answer questions in immune cell signaling, building toward the augmentation of cell-based immunotherapy. He has received several awards including a USF Faculty Outstanding Research Achievement Award, an NIGMS MIRA R35, an NSF CAREER Award and an ORAU Ralph E. Powe Junior Faculty Enhancement Award.