Inflammation is important in mediating host defense. However, chronic inflammation can lead to the development of numerous lifelong disorders. Specialized pro-resolving lipid mediators (SPMs) are lipid mediators that actively inhibit inflammation and enhance resolution. SPMs are generated through the actions of various lipid mediator biosynthetic enzymes such as 5-lipoxygenase (5LO or ALOX5). While there has been a lot of progress elucidating lipoxygenase- mediated biosynthetic pathways leading to SPM production, the upstream cues governing activation and regulation of ALOX5 are still not fully understood. Our laboratory discovered that engagement of the NOD2 pathway led to the production of not only pro-inflammatory lipid mediators, but also SPMs. In order to understand how NOD2 was mediating these effects, we investigated the role of NOD2 in influencing the post-translational modification, localization, and stabilization of ALOX5. We found that while the presence of NOD2 promoted the activating phosphorylation of ALOX5, it also decreased the levels of ALOX5 in a dose-dependent manner. Further pulse-chase analysis indicated that although NOD2 initially decreased ALOX5 protein levels, it ultimately prolonged the half-life of total and phosphorylated ALOX5. It was also observed that certain disease associated NOD2 variants caused dysregulated phosphorylation of ALOX5 or dysregulated localization of activated ALOX5. Lastly, mass spectrometry analysis of ALOX5 in M1 or M2 polarized THP-1 macrophages stimulated with the NOD2 agonist MDP indicated differential proline hydroxylation of ALOX5 under these conditions. Understanding how NOD2 influences ALOX5 may shed light on the mechanisms which are dysregulated under pathological conditions and could inform the potential utility of various inhibitors of lipid mediator production or of NOD2/RIP2 signaling for treatment of specific inflammatory diseases.
| Identifer | oai:union.ndltd.org:ucf.edu/oai:stars.library.ucf.edu:honorstheses-2718 |
| Date | 01 January 2023 |
| Creators | Uppada, Harshitha |
| Publisher | STARS |
| Source Sets | University of Central Florida |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | Honors Undergraduate Theses |
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