Biology
Permanent URI for this collectionhttps://hdl.handle.net/11274/15791
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Browsing Biology by Author "Averitt, Dayna L"
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Item Identifying and characterizing regulators of a bone morphogenetic protein cell signaling pathway in Caenorhabditis elegans(2022-12-01T06:00:00.000Z) Lakdawala, Mohammed Farhan; Gumienny, Tina L; Averitt, Dayna L; Brower, Christopher; Savage-Dunn, Cathy; Spencer, JulietCell-to-cell communication pathways ensure proper development of organisms. One group of signaling pathways that animals use is the bone morphogenetic protein (BMP) family. BMPs play significant roles in embryogenesis, development of skeletal and cardiovascular systems, among many others. Misregulation of BMP signaling leads to various diseases and thus needs to be tightly regulated. An understanding of how this pathway is itself controlled and performs diverse functions throughout the lifetime of living organisms is not very well understood. To identify new regulators of the BMP signaling pathway we used the roundworm C. elegans. DBL-1/BMP signaling is a dose dependent regulator of body size. Over expression of DBL-1 makes animal longer and suppression of the pathway reduces animal body size. Using genetic and imaging tools, we identified a group of genes called dumpy (dpy) that genetically interact with DBL-1/BMP signaling. Most of the identified dpy genes code for extracellular matrix components and/or are associated with body size regulation. We showed that loss of some dpy genes suppresses expression of the DBL-1 ligand and DBL-1 signaling activity. Using RNA sequencing, we identified genes that are differentially regulated by the DBL-1 signaling pathway; many of which regulate body size. Furthermore, we identified two dpy genes, dpy-2 and dpy-9, that have stage-specific interactions with the DBL-1 signaling pathway. Additionally, we characterized the interaction between DBL-1 pathway and dpy-24/blmp-1. BLMP-1 is a transcription regulator and controls some organismal traits that the DBL-1 pathway also affects. We showed DBL-1 signaling and BLMP-1 regulate each other. We also identified a physical interaction between BLMP-1 and the SMADs, downstream regulator of DBL-1 pathway using yeast-two hybrid system. Using bioinformatics and qPCR analysis, we showed that the interaction between SMADs and BLMP-1 is required to regulate some common downstream target genes. Lastly, we performed epistasis studies and identified that the DBL-1 signaling pathway and BLMP-1 work partially independently to regulate body size and work together to regulate brood size. Collectively, we identified additional regulators and interactors of the DBL-1 signaling pathway, and our findings provide insights about a new molecular mechanism by which DBL-1 pathway regulate certain organism traits.Item Sex differences and the effects of stress on neuroanatomical structures involved in modulating inflammatory pain(December 2023) Cantu, Daisy Jacqueline; Averitt, Dayna L; Hynds, DiAnna; Lybrand, Zane; Brower, Christopher; Anne MurphyApproximately 1 in 5 people suffer from pain worldwide, and women experience more recurrent, severe, and longer-lasting pain. Pain conditions more prevalent in women include migraine, fibromyalgia, irritable bowel syndrome, and temporomandibular joint disorder. Stress can influence pain, and females are more susceptible to stress-exacerbated orofacial pain. As preclinical studies have largely been conducted in males, the mechanisms underlying the effects of stress on pain in females are understudied. Orofacial pain is relayed by sensory neurons in the trigeminal ganglia (TG) that synapse in the brainstem trigeminal nuclear complex (TnC). Ascending pain pathways relay pain signals to the cortex and can initiate descending pain modulation via emotive nuclei in the amygdala and hypothalamus that project to the periaqueductal gray. Our overarching hypothesis is that stress exacerbates orofacial pain to a greater degree in female rats, and there are sex differences in the trigeminal neurocircuitry that transmit somatosensory information to the brain. We utilized behavioral, neuroanatomical, pharmacological, and transcriptomic approaches to investigate sex differences in the effects of stress in a rat model of orofacial pain. We report that stress exacerbates orofacial pain to a greater degree in female rats. Neuroanatomical analyses of the TG and TnC implicate a sex-specific pain mechanism whereby females have dampened GABAergic neurotransmission during stress-exacerbated orofacial pain. In support, pain behaviors were reduced by treatment with a GABAB receptor agonist. Further, when we explored the pain-modulatory projections of the amygdala and hypothalamus to the periaqueductal gray, we uncovered sex differences in their organization and activation by pain, supporting a sexually dimorphic role of emotive brain areas in modulating pain. Overall, our data indicate that stress differentially alters sensory neuron input to the brainstem and is subject to sex-specific descending modulation from emotive centers in the brain. We postulate that women experiencing stress-exacerbated orofacial pain would benefit from a pain management regimen that includes a pharmacological intervention to boost GABA signaling concomitant with interventions that manage stress. Lastly, our transcriptomics data may indicate novel pain therapeutic targets related to the extracellular matrix, which may contribute to the effects of stress on pain chronification in the trigeminal system.Item The functional characterization of a novel protein, Liat1.(2022-12-01T06:00:00.000Z) Arva, Akshaya; Brower, Christopher; Averitt, Dayna L; Conrad-Webb, Heather; Juliet V Spencer; Douglas Root; Averitt, Dayna LThe Ligand of Ate1 (Liat1) was discovered in our lab by its interaction with Arginyl tRNA-protein transferase-1 (Ate1), a component of the N-degron pathway of protein degradation. However, its functional significance remains unknown. This study is the first functional characterization of Liat1. Here, we found that the N-terminal half of Liat1 is intrinsically disordered. Intrinsically disordered proteins tend to drive a phenomenon called liquid-liquid phase separation (LLPS). This involves the reversible conversion of a homogenous solution into separate phases of distinct density through the condensation of one or more of its biomolecular components. This process results in the formation of membrane-lacking subcellular organelles that organize cellular components and enhance biochemical reactions. Using a combination of immunocytochemistry, bimolecular fluorescence complementation, and biochemical techniques, we found that Liat1 undergoes LLPS in the dense fibrillar compartment (DFC) of the nucleolus. Mutational analysis revealed that a low complexity, poly-lysine region contained in the N-terminal intrinsically disordered region of Liat1 is required for its self-interaction and nucleolar phase separation. In addition to Ate1, Liat1 interacts with Jmjd6, a bifunctional enzyme with demethylase and lysl-hydroxylase activity. We show that Jmjd6 modifies Liat1 and prevents its nucleolar targeting through in a manner that requires its hydroxylation activity. In efforts to identify the Liat1 amino acids modified by Jmjd6, we expressed Jmjd6 in bacterial cells and established an in vitro assay to monitor it lysyl-hydroxylation under purified conditions. In efforts to characterize the nucleolar function of Liat1, we monitored its behavior in response to specific stress conditions. Specifically we found that Liat1 remains in the core of the nucleolus when rDNA transcription is inhibited; conditions which disrupt the overall structure of the nucleolus. Furthermore, we found that Liat1 is upregulated and localized to the cytosol during viral infection. These studies reveal distinct behaviors of Liat1 in response to specific forms of cell stress. Finally we found that Liat1 interacts with small ribosomal subunit proteins and that cells lacking Liat1 have defects in rRNA processing. Collectively, our data indicates that Liat1 participates in LLPS in the dense fibrillary component of the nucleolus in a manner regulated by Jmjd6; and suggest that it plays a significant role in rRNA processing and/or assembly as well as in various cellular stress responses.Item The role of Arginyltransferase 1 in body weight homeostasis(August 2023) Alkhatatbeh, Mosleh Ahmed 1977-; Brower, Christopher; Conrad-Webb, Heather; Mills, Nathaniel; Averitt, Dayna L; Na, ElisaPrevious studies reported that systemic deletion of Arginyltransferase 1 (ATE1) in mice results in dramatic fat loss and resistance to diet-induced obesity. However, the mechanisms through which ATE1 influence energy metabolism remain unclear. Here, we investigated the hypothalamic role of ATE1 by examining the effects of the anorectic hormone leptin in wild type (WT) and ATE1-knockout (ATE1-KO) mice maintained on a normal chow diet, or a high fat diet. We found that on both diets, ATE1-KO mice weighed significantly less than WT mice despite consuming more food (hyperphagia). We also found that, similar to WT mice on normal chow, ATE1-KO mice remain responsive to leptin on NC. However, ATE1-KO mice had significantly lower circulating plasma leptin than wild type mice on both diets. This explains (at least in part) their hyperphagia. Interestingly, we also found that leptin reduces the expression of ATE1 in the hypothalamus, indicating that decreased ATE1 activity may be a component of the Leptin regulatory network driving a lean phenotype. In contrast to normal chow, we found that ATE1-KO mice became leptin resistant on high fat diet. Remarkably, we found that the loss of Ate1 gene function reverses high fat diet-induced obesity. Collectively, these results strongly suggest that the inhibition of ATE1 may offer an effective therapeutic strategy to combat obesity.