Through the Life-Health Sciences Internship program, I have been blessed with the opportunity to conduct paid undergraduate research through the Indiana University School of Medicine. After a multi-interview process, I was placed with Dr. Gustavo Arrizabalaga in the Department of Pharmacology and Toxicology. His lab, along with a few others, works to study the parasite Toxoplasma gondii. This parasite is present worldwide, can spread to any warm-blooded animal, and affects nearly a third of the world's human population. Healthy individuals exhibit flu-like symptoms when they are infected; however, fetuses, immunocompromised individuals, and organ transplant patients infected with the parasite may develop congenital deformities and toxoplasma encephalitis. In the United States, Toxoplasma is the leading cause of death by foodborne illness. In order to understand the parasite and hopefully one day develop anti-parasitic drugs, the lab is working to study Toxoplasma's cell-signaling pathways, since leaving the host cell (a process known as egress) is vital for Toxoplasma survival and proliferation.
As an undergraduate research student, I am working under Dr. Chunlin Yang, PhD, to learn about lab practices, techniques, and processes. It is my goal to 1) demonstrate mastery of multiple laboratory techniques in both application and understanding, 2) evaluate various health career goals and prepare for success in the field by developing transferrable, career-related, and presentation skills, and 3) relate my project to the larger implications of work being done to further understand the experimentation of Toxoplasma.
I have thoroughly enjoyed my experience thus far in this laboratory. I have been able to develop scientific research skills as well as improved critical thinking skills. Much of the material I have encountered has related to my academic studies. This semester, I was surprised to learn information in my Biochemistry course that coincides with skills and information I have been taught in the lab! My mentors have been extremely benefical to my learning process in that they do their best to allow me to learn the background information relating to individual experiments. They also encourage me to make mistakes and learn from them, which ultimately has improved my techniques and critical thinking skills. I avoid making the same mistake twice because I remember what it felt like to be responsible for an error in an experiment.
ABSTRACT
Localization investigation of the potential parasitophorous vacuole or plasma membrane localized PP2Cs in Toxoplasma gondii
Emily M. Sampson1, Chunlin Yang1, and Gustavo A. Arrizabalaga1,2
1Department of Pharmacology and Toxicology, Indiana University School of Medicine; 2Department of Microbiology and Immunology, Indiana University School of Medicine
Toxoplasma gondii is an obligate intracellular parasite that can invade nearly any nucleated cell of warm-blooded animals, including human beings. The infection of T. gondii usually will not show serious symptoms in immune-competent human hosts because the actively multiplying tachyzoites of this parasite will switch into slowly dividing bradyzoites enclosed in tissue cysts under the pressure of the host immune system. Conversely, for those immunocompromised or immunosuppressed individuals, incompetent immune systems may not pressure the parasites enough to switch forms. Consequently, repeating lytic cycles of the tachyzoites will cause widespread tissue damage, leading to the life threating disease toxoplasmosis; furthermore, the available therapeutic drugs are ineffective in many toxoplasmosis patients. As the main contributor of the disease, the lytic cycle of T. gondii has been extensively investigated to determine the regulation of each step, including host cell attachment, invasion, parasitophorous vacuole (PV) formation, replication, and egress. Previous studies have reported that phosphorylation is key in the regulation of the T. gondii lytic cycle, especially in regards to the central roles of apicomplexan-specific calcium dependent protein kinases (CDPKs). Contrarily, no protein phosphatases have been identified to be involved; therefore, identifying the protein phosphatase(s) which serve(s) as the regulator of the T. gondii lytic cycle is the primary interest of this study, with the focus on those potentially localized in the plasma membrane and PV. The proteins which are secreted into the PV usually contain signal peptides, and those localized in the plasma membrane commonly possess transmembrane domains. Thus, we selected two signal-peptide containing (TgGT1_270320, TgGT1_278510) and one transmembrane domain containing (TgGT1_304955) protein phosphatases in this study. Also, we included another protein phosphatase (TgGT1_243990), whose homolog in budding yeast localizes in the plasma membrane, although it does not contain any transmembrane domains. Briefly, we endogenously tagged these four genes with a triple HA tag, and are conducting immunofluorescence assays (IFAs) to determine their localizations.
Mentors: Gustavo A. Arrizabalaga, Department of Pharmacology and Toxicology and Department of Microbiology and Immunology, IU School of Medicine; Chunlin Yang, Department of Pharmacology and Toxicology, IU School of Medicine; Brandi Gilbert, Life Health Sciences Internship Program, IUPUI.
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