Impact of nuclear envelope protein degradation on neutrophil lipid content and migration

Project Summary & Abstract Neutrophils are the front-line defenders in innate immunity and are the most abundant circulating immune cells in the human body. Unfortunately, neutrophils are also major players in perpetuating autoinflammatory conditions. Neutrophils possess remarkable nuclear deformability, a necessary requisite for their migration from blood vessels through micrometer sized intercellular spaces towards sites of inflammation. The overarching goal of this proposal is to design new approaches to control neutrophil migration by manipulating nuclear deformability. Preliminary work has shown 69 differentially regulated nuclear envelope genes associated with neutrophil differentiation. Many of these are associated with the nuclear lamina and lipid homeostasis proteins. This proposal aims to use a nanobody-based protein degradation system developed by the Dassama lab, called NanoBridge, to selectively degrade these differentially produced proteins. It proposes to use a combination of microscopy and lipidomics to assess the impact of target degradation on nuclear lipid composition and how lipid composition influences neutrophil nuclear deformability. Novel aspects of this proposal include 1) development of a system for the targeted degradation of nuclear envelope proteins, 2) assessment of lipid content in the neutrophil nucleus following targeted degradation of upregulated lipid homeostasis proteins, 3) determination of neutrophil morphology and migration following targeted degradation of understudied nuclear envelope proteins and 4) development of a system to modulate neutrophil migration via manipulation of its nuclear deformability by targeted protein degradation. The project goals will allow me to develop a well-rounded skillset in cell biology and targeted protein degradation. I will further have an opportunity to gain experience in lipidomics technology. These skills will combine with my pre-existing experience in natural product biosynthesis from my PhD work to give me a well- rounded background that will facilitate my independent research career.