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Identification and Molecular Characterization of Zebrafish (Danio rerio) Leukocyte Immune-Type Receptors (DrLITRs)
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- Author / Creator
- Gurupalli, Hima V
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In response to foreign invaders, innate immune cells use cell surface-expressed receptors to trigger intracellular signalling cascades that control various effector responses designed to destroy microbes. These specialized receptors are known as immunoregulatory receptors and structurally they exist as stimulatory and inhibitory types. In channel catfish (Ictalurus punctatus), the leukocyte immune-type receptor (IpLITR) family consists of multiple members, each with variable signalling abilities. Using in vitro approaches, our previous work has shown that IpLITRs also function as potent regulators of antimicrobial responses including phagocytosis, degranulation, cytokine secretion and cytotoxicity. To better understand the potential roles of LITRs in vivo, my thesis research focused on using zebrafish as a model organism to further examine teleost LITRs. I cloned and sequenced four putative Danio rerio (Dr)LITRs, termed DrLITR 1.1, DrLITR 1.2, DrLITR 15.1 and DrLITR 23.1 that were identified in the zebrafish genome. These receptors are related to IpLITRs and they are also distantly related to important immunoregulatory receptors found in mammals (e.g.FcRLs, SIGLECs, and CEACAMs). Structurally, DrLITRs 1.1 and 1.2 contain three extracellular immunoglobulin (Ig) domains, a transmembrane (TM) region and a cytoplasmic tail (CYT) containing both immunoreceptor tyrosine-based activation motif (ITAM) and immunoreceptor tyrosine-based inhibition motif (ITIM). DrLITR 15.1 contains four Ig domains, a TM segment and a CYT region with two ITIMs and one immunoreceptor tyrosine-based switch motif (ITSM). DrLITR 23.1 contains six Ig domains, a positively-charged TM segment and a CYT region devoid of any recognizable tyrosine-based motifs. I also examined the expression of DrLITRs throughout ontogeny and adulthood with and without an immunostimulant. These results showed that all four DrLITR-types are expressed as early as 1 hour post fertilization (hpf) and they remain present throughout embryonic development and adulthood. Although the pro-inflammatory cytokine IL1β displayed significant increases in expression at 3, 6, 8 and 12 hours using a visceral cavity based inflammation assay, the expression profiles of DrLITR 1.1, DrLITR 1.2, DrLITR 15.1 and DrLITR 23.1 displayed different trends over the same period following an intraperitoneal injection of zebrafish with 1 µg/mL zymosan. DrLITR 1.1 was significantly upregulated at 12 hours after zymosan exposure while DrLITR 1.2 was significantly upregulated at 3, 6 and 12 hours post zymosan injection. DrLITR 15.1 displayed no statistically significant upregulation/downregulation patterns at any time point while DrLITR 23.1 was downregulated at 24 hours after zymosan injection. Overall, this work sets the stage for establishing zebrafish as a model system to study these novel immunoregulatory receptors during various developmental stages of fish and during pathogen-induced inflammatory responses.
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- Subjects / Keywords
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- Graduation date
- Spring 2020
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- Type of Item
- Thesis
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- Degree
- Master of Science
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- License
- Permission is hereby granted to the University of Alberta Libraries to reproduce single copies of this thesis and to lend or sell such copies for private, scholarly or scientific research purposes only. Where the thesis is converted to, or otherwise made available in digital form, the University of Alberta will advise potential users of the thesis of these terms. The author reserves all other publication and other rights in association with the copyright in the thesis and, except as herein before provided, neither the thesis nor any substantial portion thereof may be printed or otherwise reproduced in any material form whatsoever without the author's prior written permission.