Microbiology and Immunology Department Theses and Dissertations

Permanent URI for this collection

https://hdl.handle.net/1805/1660

Find out more about this Program at: http://micro.medicine.iu.edu

Browse

Now showing 1 - 5 of 5

Characterization of a Novel Fis1 Interactor Required for Peripheral Distribution of the Mitochondrion of Toxoplasma Gondii
(2021-02) Jacobs, Kylie; Arrizabalaga, Gustavo; Gilk, Stacey; Graham, Brett; John, Chandy; Yang, Frank
Toxoplasma’s singular mitochondrion is extremely dynamic and undergoes morphological changes throughout the parasite’s life cycle. While intracellular‚ the mitochondrion is maintained in a lasso shape that stretches around the parasite periphery and is in close proximity to the pellicle‚ suggesting the presence of membrane contact sites. Upon egress‚ these contact sites disappear‚ and the mitochondrion retracts and collapses towards the apical end of the parasite. Once reinvaded‚ the lasso shape is quickly reformed‚ indicating that dynamic membrane contact sites regulate the positioning of the mitochondrion. We discovered a novel protein (TgGT1_265180) that associates with the mitochondrion via interactions with the fission related protein Fis1. Knockout of TgGT1_265180‚ which we have dubbed LMF1 for Lasso Maintenance Factor 1‚ results in a complete disruption of the normal mitochondrial morphology. In intracellular LMF1 knockout parasites, the mitochondrial lasso shape is disrupted‚ and instead it is collapsed as normally only seen in extracellular parasites. Additionally, proper mitochondrial segregation is disrupted‚ resulting in parasites with no mitochondrion and extra mitochondrial material outside of the parasites. These gross morphological changes are associated with a significant reduction of parasite propagation and can be rescued by reintroduction of a wildtype copy of LMF1. Co-immunoprecipitations and Yeast Two-Hybrid predict interactions with the parasite pellicle. Therefore, we hypothesize that LMF1 mediates contact between the mitochondrion and the pellicle in a regulatable fashion‚ and that the LMF1-dependent morphodynamics are critical for parasite propagation. Current studies are focused on characterizing the consequences of mitochondrial collapse and identifying proteins that interact with LMF1 to position the mitochondrion to the periphery of the parasite.
Guanabenz Reduces Hyperactivity and Neuroinflammation Caused by Latent Toxoplasmosis in Mice
(2020-02) Martynowicz, Jennifer Marie; Sullivan, William J., Jr.; Arrizabalaga, Gustavo; Boehm II, Stephen L.; Gilk, Stacey D.; Spinola, Stanley M.
Toxoplasma gondii is an intracellular parasite that causes persistent, lifelong infection in one-third of humans worldwide. The parasite converts from a lytic, actively replicating form (tachyzoite) into a latent tissue cyst form (bradyzoite) that evades host immunity and is impervious to current drugs. While acute infection can be life threatening to immunosuppressed individuals, chronic infection has been linked to behavioral changes in rodents and neurological disease in humans. Notably, chronic infection in mice leads to hyperactivity in an open field. Whether these behavioral changes are due to parasite manipulation of the host or the host response to infection remains an outstanding question. We have previously shown that the anti-hypertensive drug guanabenz reduces Toxoplasma cyst burden in the brains of BALB/c mice, providing a means to examine whether brain cyst depletion reverses behavioral changes. We used two mouse strains (BALB/c and C57BL/6) differing in their susceptibility to infection. Following drug treatment of chronically infected mice, locomotor activity in an open field was assessed. In both mouse strains, the increased hyperactivity seen during chronic infection returned to normal levels following guanabenz treatment. Guanabenz reduced brain cyst burden ~70% in BALB/c mice as expected, but it increased cyst burden 49% in C57BL/6 mice. Examination of the brains showed that guanabenz decreased inflammation and perivascular cuffing in both infected mouse strains. Our study shows for the first time that it is possible to reverse a key behavioral change associated with chronic Toxoplasma infection. Surprisingly, the rescue from parasite-induced hyperactivity correlates with a decrease in neuroinflammation instead of cyst counts, suggesting that some behavioral changes arise from host responses to infection rather than a parasite-driven process.
Inflammation by Toxoplasma gondii Infection Induces Prostatic Hyperplasia and Accompanying Urinary Dysfunction
(2024-08) Stanczak, Emily F.; Arrizabalaga, Gustavo; Bauer, Margaret; Jerde, Travis; Nakshatri, Harikrishna
Benign prostatic hyperplasia is the non-cancerous enlargement of the prostate. This syndrome develops as men age and affects 50% of men by the age of 50 and 80% of men by the age of 80. BPH is associated with a pattern of symptoms and pathology in patients that can be painful, problematic, and lower the quality of life. BPH is a multifactorial disease which may develop due to lifestyle choices, genetics, metabolic disorder, and potentially infection of the organ. Interestingly, previous work from our research group showed that the common parasite T. gondii can infect and establish a chronic infection in the prostate of mice resulting in histological hyperplasia and glandular nodule formation, reminiscent of that observed in men with BPH. This leads to the hypothesis that T. gondii contributes to BPH in humans. In this work, we investigated whether there is a correlation between T. gondii infection and BPH using a cohort of age matched BPH diagnosed and non-BPH diagnosed control donors. My data show that men diagnosed with BPH are significantly more likely to have evidence of T. gondii infection than undiagnosed controls. Additionally, men with antibodies against T. gondii and BPH had significantly more severe pathology in several categories including and most notably in epithelial and glandular nodules seen only in a steroid hormone model of the disease. Moreover, I have determined that mice infected with parasites demonstrated abnormal urination pattern behavior indicating lower urinary tract dysfunction. Based on these results, we conclude that T. gondii can contribute to the development and severity of BPH and BPH pathology in humans. In addition, T. gondii can potentially be used as a model for BPH and BPH-like symptoms and pathology in mice and other model organisms.
Investigating the Function and Therapeutic Potential of the GCN5b Bromodomain in Toxoplasma Gondii
(2020-06) Hanquier, Jocelyne Nicole; Sullivan, William J., Jr.; Arrizabalaga, Gustavo; Gilk, Stacey
The obligate intracellular protozoan parasite Toxoplasma gondii is a medically relevant pathogen that has infected a third of the world’s population. Toxoplasma is the causative agent of toxoplasmosis, which can have severe repercussions such as encephalitis and even death in immunocompromised patients. Current treatments for toxoplasmosis only target acute infection and can be toxic to patients, resulting in complications including allergy and bone marrow suppression. Thus, the identification of novel drug targets and therapies for toxoplasmosis is vital. Epigenetic modulators of lysine acetylation, including ‘writers,’ ‘erasers,’ and ‘readers,’ have been identified as promising drug targets for protozoan parasites. The lysine acetyltransferase (KAT) GCN5b appears to be an essential gene for Toxoplasma viability. The KAT domain of GCN5b is essential to GCN5b function and is targetable by small molecule inhibitors. While the acetyltransferase activity of this gene is well-characterized, the functionality of its C-terminal bromodomain (BRD) remains to be understood. Bromodomains are readers of lysine acetylation, and recently, bromodomain inhibitors have shown promise in a number of human diseases, as well as in protozoan parasites. We hypothesized that the GCN5b bromodomain is critical for Toxoplasma viability. The data reported herein suggest that the GCN5b bromodomain is important for tachyzoite viability and may serve as a novel therapeutic target in Toxoplasma.
The Role of Mammalian Lipid Transport Protein ORP1 During Coxiella Burnetii Infection
(2022-05) Schuler, Baleigh Elizabeth; Gilk, Stacey D.; Arrizabalaga, Gustavo; Spinola, Stanley; Harrington, Maureen; Day, Richard
Coxiella burnetii is an intracellular bacterium that causes the human disease Q fever. C. burnetii is transmitted from infected animals to humans through inhalation of infectious droplets. Acute Q fever is a flu-like illness lasting 10-14 days. Patients often have respiratory symptoms and present with pneumonia. Patients with suppressed immune systems or valvular heart disease can develop chronic Q fever, which causes endocarditis and vasculitis long after initial infection. Chronic Q fever is difficult to treat, and if untreated, is typically fatal. Currently, the United States lacks any vaccine for Q fever. In order to better prevent and treat this disease, it is important to understand how C. burnetii interacts with mammalian cells. Within the host cell, C. burnetii forms a large, acidic Coxiella-containing vacuole (CCV) and uses a Type 4B secretion system (T4SS) to secrete effector proteins into the host cell cytoplasm. While the CCV membrane is rich in sterols, cholesterol accumulation in the CCV is bacteriolytic, suggesting that C. burnetii regulation of lipid transport is critical for infection. The mammalian lipid transport protein ORP1L localizes to the CCV membrane and mediates CCV-ER membrane contact sites. ORP1L functions in lipid transport, including cholesterol efflux from late endosomes/lysosomes. Its sister isoform ORP1S binds cholesterol but localizes to the cytoplasm and nucleus. In ORP1- null cells, we found that CCVs were smaller than in wildtype cells, highlighting the importance of ORP1 in CCV development. CCVs in ORP1-null cells had higher cholesterol content than CCVs in wildtype cells, suggesting ORP1 functions in cholesterol efflux from the CCV. ORP1-null MH-S cells do not accumulate lipid droplets upon C. burnetii infection, supporting our hypothesis that ORP1 promotes cholesterol transfer from the CCV to the ER, as lipid droplets form from neutral lipids in the ER. While the absence of ORP1 led to a C. burnetii growth defect in MH-S cells, there was no growth defect in HeLa cells. Together, our data demonstrate that C. burnetii uses the host sterol transport protein ORP1 to promote CCV development, potentially by using ORP1 to facilitate cholesterol efflux from the CCV to diminish the bacteriolytic effects of cholesterol.

Browse

Browsing Microbiology and Immunology Department Theses and Dissertations by Author "Arrizabalaga, Gustavo"

Results Per Page

Sort Options