Date of Award

5-2019

Document Type

Thesis

Degree Name

Master of Science (MS)

Department

Biological Sciences

Committee Member

Lesly A Temesvari, Committee Chair

Committee Member

Zhicheng Dou

Committee Member

Lukasz Kozubowksi

Abstract

Entamoeba histolytica is an intestinal parasite infecting over 50 million people worldwide and is the causative agent of amebic dysentery and amoebic liver abscess. In the human host and nonhuman primates, E. histolytica experiences stress brought on by nutrient deprivation and the host immune response. To be a successful parasite, E. histolytica must counter the stress; therefore, understanding the stress response may uncover new drug targets. In many systems, the stress response includes down-regulation of general protein translation, which is regulated by phosphorylation of eukaryotic initiation factor (eIF-2α). Previous work in E. histolytica has demonstrated that EheIF-2α phosphorylation increases significantly when exposed to long-term serum starvation, oxidative stress, and long-term heat shock. However, the effects of nitrosative and endoplasmic reticulum (ER) stresses, on the eIF2α protein translation control system have yet to be evaluated. Nitrosative stress is part of the host’s immune response and ER stress can be caused by several physiological or pathological factors. We treated E. histolytica cells with different reagents to induce nitrosative stress (DPTA-NONOate and SNP) or ER stress (BFA and DTT). We examined the morphology of the ER, tracked phosphorylation of eIF2α, and assessed protein translation in the control and stressed cells. While all four stress-inducing reagents caused a global reduction in protein translation, only DTT was capable of also inducing changes in the morphology of the ER (consistent with ER stress) and phosphorylation of EheIF-2α. This suggests that DTT authentically induces ER stress in E. histolytica and that this stress is managed by the eIF2α-based system. This was supported by the observation that cells expressing a non-phosphorylatable version of eIF2α were also highly sensitive to DTT-stress. Since protein translation decreased in the absence of phosphorylation of eIF2α (after treatment with DPTA-NONOate, SNP or BFA), the data also indicate that there are alternative protein-translational control pathways in E. histolytica. Overall, our study further illuminates the nitrosative and ER stress responses in E. histolytica.

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