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ABSTRACT KORTUM, AMANDA NOELE. Comparative Analysis Of ABSTRACT KORTUM, AMANDA NOELE. Comparative Analysis of Tripartite Motif Containing 9 Protein Expression in Macrophages and Neutrophils. (Under the direction of Dr. Jeffrey Yoder). The persistence of an immune response can contribute to the development of disease states, increasing the extent of tissue damage or perpetuating the disease condition. Chronic inflammation is often characterized by high numbers of macrophages and neutrophils (phagocytes) that release ROS, cytotoxic factors, and cytokines, which can further amplify inflammation. Phagocytes have been shown to contribute to disease in both natural and laboratory-induced models of chronic inflammation in veterinary species. Understanding the cellular mechanisms that mediate phagocyte function may reveal how these cells could be therapeutically targeted during persistent inflammation. The E3 ubiquitin ligase TRIM9 is highly expressed in the brain where it mediates axon migration and synaptic vesicle release. While current literature on TRIM9 function in the brain is limited to human cell lines and mouse models, a recent study demonstrated TRIM9 is critical to normal macrophage cell shape and motility in zebrafish. In this study, I aimed to establish a novel mammalian model for the study of TRIM9 in phagocytes. Through the comparative analysis of the TRIM9 expression in macrophages and neutrophils from human, horse, pig, dog, and mouse, I identify TRIM9 isoforms exhibit tissue specificity that are observed across species. Furthermore, based on similar tissue expression of TRIM9 in humans and horses, I propose the use of an equine model for future studies on TRIM9 function in phagocytes. © Copyright 2019 by Amanda Noele Kortum All Rights Reserved Comparative Analysis of Tripartite Motif Containing 9 Protein Expression in Macrophages and Neutrophils. by Amanda Noele Kortum A dissertation submitted to the Graduate Faculty of North Carolina State University in partial fulfillment of the requirements for the degree of Doctor of Philosophy Comparative Biomedical Sciences Raleigh, North Carolina 2019 APPROVED BY: _______________________________ _______________________________ Dr. Jeffrey Yoder Dr. Samuel Jones Committee Chair _______________________________ _______________________________ Dr. Barbara Sherry Dr. David Muddiman DEDICATION To my Great-Grandma Georgie, who started my education early in life with rag-bag-sag-nag and my „arithmetic‟ lessons and My Grandma TheaLou, who never hesitated putting dinner on the table two hours late when a little girl butchering chickens asked “What‟s this?” ii BIOGRAPHY Amanda grew up in the rural town of Ekalaka, Montana. This was 100-miles-from-a- McDonald‟s, population 300, kind of rural. She spent much of her childhood on her grandparents‟ ranch. From watching a caesarian section on a pregnant cow in her Grandpa‟s barn to having ducks, rabbits, guinea pigs, dogs, cats, goats, and ferrets as pets, Amanda‟s passion for animals blossomed early. It wasn‟t until high school that a combined interest in biology, and specifically biological research, drew Amanda away from her original career goal of psychiatry. As the only female in a small Science Research class, Amanda learned the scientific method, developing her own hypotheses, creating experiments and analyzing the results to make conclusions, all while putting up with the antics of her esteemed colleagues, Reed (now Dr. Lambert), Eric (newspaper editor), and Nathan (almost Dr. Carroll). During college at the University of Montana Western, Amanda pursued scientific research, working in the microbiology lab of her undergraduate mentor, Dr. Mike Morrow. Through four years of research on Candida albicans, the pathogenic yeast, Amanda found her true passion in life, infectious disease. After graduating with a degree in Biology, Amanda went on to put her degree to work, building up her resume before applying to veterinary school. A year spent as a Scientific Researcher I, at the University of Washington School of Medicine only reassured her that pursuit of dual DVM and PhD degrees was the correct path for her. Through the culture shock of a big city and the concrete of Seattle, WA, Amanda chose to attend North Carolina State University College of Veterinary Medicine after many stressful interviews. It was also, coincidently, the choice of her future DVM/PhD/soulmate and partner-in-crime, Hannah Reynolds. iii Amanda joined Dr. Jeff Yoder‟s Comparative Immunology Lab due to a combined interest in the cat vaccines and the glowing zebrafish Jeff promised, but largely due to Jeff‟s passion and excitement for his research. It was only after joining the lab that Amanda realized comparative immunology was the bridge that connected her goal of helping animals through veterinary medicine to her passion for research, scientist-minded brain, and perverse love of infectious disease. Since beginning the DVM/PhD program, Amanda has had the privilege of learning veterinary medicine with 100+ now DVMs, she has seen graduate student colleagues become doctors and doctor doctors, and she met and married her husband, Josh. As a fellow in the USDA APHIS NBAF Scientist Training Program, Amanda will fulfill her longtime goal of working for the USDA Foreign Animal Disease and Diagnostic Laboratories at Plum Island, NY upon completion of DVM/PhD program. After the completion of the National Bio and Agro-Defense Facility, Amanda will move to Manhattan, KS to finish her service commitment to the USDA. iv ACKNOWLEDGMENTS My journey to this point has been filled with so many supportive and loving individuals; the list could be longer than my dissertation. First and foremost, I need to thank my parents for their unrequited belief in me, my dreams, and the journey it has taken me to get where I am today. My upbringing has prepared me for many of the challenges I have faced and have waiting for me in my future, and for that, I am eternally grateful. Likewise, I have numerous other family members that have offered their unconditional support to me in the last thirty years. I am so thankful for my grandfather and uncle, my brother and his wife, my sister Becky and her family- all people that have flown/driven me places and sent words and gifts of encouragement. My husband, my patient, kind, creative, intelligent husband, Josh- you have seen me at my utter worst and I hope, what I consider my very best. You don‟t count as people, because when I am all peopled out, and need to be alone, you are the only one I want with me. You are everything I could have ever wanted in a life partner and your selflessness, encouragement, and love have fueled these last few long years. Countless friends deserve thanks: my childhood besties, Jen, Cecile, and Blake, my college pal, Trish, my friend forever in education with me, Hannah. You all build me up, in your own ways and are always there for me. Hannah, obviously I couldn‟t have gotten married without you, but I also could not have survived these last 7 years without you. From vet school, Alex Davis (and Syd) are the friends I didn‟t know I wanted, and you always know how to make me laugh. To the DVM/PhD crew, I truly believe one of the main reasons any of us have finished is that we are all in this together. The other reason is Sam Jones. Obviously. To Emily especially, you are one of the most positive, brilliant people I know and your inappropriate use of „that‟s cute‟ has made me smile many times over the years. v My labmates deserve a large round of acknowledgement. From putting up with disruptive chitchat to answering questions and only rolling your eyes 50% of the time, Dustin- I‟m still cranky you finished before me. Jess, you are a role model in everything you do, vet school, research, motherhood, and being an overall amazing, kind person. Deb, Ashley, Hayley, and Ivan answered every question patiently, while Drake, Joy, Thomas, Emma, and Kathryn put up with obnoxious requests. Shannon Chiera, Alix Berglund, Caroline Johnson, and the Sheets and Jones Labs offered their expertise, samples, and encouragement. I have to tribute my love of biology and research to my high school teachers, Mrs. Frye and Mrs. Carroll. You fostered my curiosity and opened my eyes to the scientific method. My undergraduate mentor, Dr. Michael Morrow, you made me realize that I thought like a scientist, that I would always have questions, and a PhD would give me the tools to answer those questions. You also exposed me to microbiology and from that, stemmed my interest in infectious disease. Dr. Mike, you always supported my goals, through words of encouragement, letters of recommendation, and pushes to apply for opportunities that opened doors, none of which ended when I left UMW. Lastly, I would like to thank my committee, Drs. Barb Sherry, Sam Jones, and Dave Muddiman. You are all experienced, skilled, and proven in your field and I am eternally grateful for not only your scientific expertise, but your guidance towards my development as a scientist and my future career. To Dr. Jeff Yoder, I have nothing but appreciation for your support. I was never pressured to answer a scientific question that didn‟t interest me, I always felt I could approach you with strange questions or goofy suggestions, even the BFP+ bacteria, and you allowed me to take my own path through my PhD training to gain experiences that would put me closer to my dream career. vi TABLE OF CONTENTS LIST OF TABLES ...................................................................................................................... viii LIST
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