Browsing by Author "Flak, Jonathan"

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    Investigation of the Knockout of LMF1 on the Transcriptome of Toxoplasma gondii
    (2024-01) Thibodeau, Katherine E.; Arrizabalaga, Gustavo; Absalon, Sabrina; Fehrenbacher, Jill; Flak, Jonathan; Schmidt, Nathan
    Toxoplasma gondii is an obligate intracellular apicomplexan parasite that infects one third of the global population. There are limited treatments for Toxoplasmosis, however a potential drug target for Toxoplasma is its mitochondrion. While much is known about the function of this organelle in Toxoplasma, little is known about the mechanisms that regulate mitochondrial structure and division. The shape of the mitochondrion changes throughout the life cycle of the parasite. When inside a host cell, the mitochondrion is in a lasso shape, stretching around the periphery of the parasite, while in extracellular parasites it is collapsed towards the apical end of the parasite. While in a lasso shape the mitochondrion shows areas of contact with the parasite pellicle. We have determined that the proteins LMF1 (associated with the outer mitochondrial membrane) and IMC10 (inner membrane complex) interact and form a reversible tether that maintains the lasso shape of the mitochondrion. When either of these proteins are knocked out, the mitochondrion collapses. To elucidate the biological relevance of the interaction between the mitochondrion and the pellicle we explored the consequence of disrupting the interaction on the transcriptome of the parasite. RNA sequencing of the LMF1 knockout strain showed a disruption in the expression of genes involved in nucleotide metabolism and Coenzyme A biosynthesis, which might be an adaptation mechanism to the disruption of mitochondrial morphology. Current work focuses on investigating the connection between mitochondrial tethering and these pathways as well as a potential role for the mitochondrion/pellicle connection in metabolite transport.
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    The Role of ABI3 in Obesity and Metabolic Regulation
    (2024-04) Smith, Daniel Curtis; Oblak, Adrian; Kim, Jungsu; Flak, Jonathan; Lasagna-Reeves, Cristian; Evans-Molina, Carmella
    Abelson Interactor Protein 3 is an adaptor protein involved in cytoskeletal remodeling. ABI3 is predominantly expressed within mononuclear phagocytotic immune cells within the brain, such as macrophages, peripherally, and microglia. Until recently, little was known about the function of the ABI3 protein, and even less was known regarding its role in disease. Following the identification of a rare mutation within ABI3 that increases the risk of developing Alzheimer’s disease, our laboratory began to investigate the impact of deleting Abi3 in mouse models. While we initially set out to investigate ABI3 in the context of neurodegeneration, we unexpectedly discovered that loss of Abi3 led to obesity in mice. This discovery and the subsequent efforts to uncover the mechanisms by which loss of Abi3 induces obesity are the subject of this dissertation. First, we demonstrate that deletion of Abi3 leads to severe obesity in aged mice. We identified significant Abi3-dependent transcriptomic changes within the hypothalamus, but not adipose tissue, of these mice. These changes occurred within pathways related to immune function, and subsequent immunostaining revealed decreased microglia number and area within the mediobasal hypothalamus of Abi3-/- mice. Next, we performed a longitudinal high-fat diet study to explore the impact of loss of Abi3 on mouse body weight and metabolic regulation during chronic nutrient excess and control conditions. Intriguingly, we found that only female Abi3-/- mice exhibited increased body weight during high-fat diet feeding. Subsequent transcriptomics from the hypothalamus of female Abi3+/+ and Abi3-/- mice from both high-fat and control diet groups revealed cytoskeletal-related changes only in the obese, high-fat diet-fed female Abi3-/- mice. Follow-up immunostaining revealed decreased microglia coverage within the mediobasal hypothalamus of the obese, high-fat diet-fed female Abi3-/- mice. While much remains to be explored regarding the precise role of ABI3 in the setting of energy balance regulation and obesity, our investigations revealed that loss of ABI3 is sufficient to induce obesity and appears to occur through altered microglia function within the hypothalamus. This dissertation represents a critical first step in the investigation of a novel regulator of obesity pathology.