Date of Award

8-2013

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Legacy Department

Biochemistry and Molecular Biology

Committee Chair/Advisor

Temesvari, Lesly A

Committee Member

Morris , James C

Committee Member

Smith , Kerry S

Committee Member

Feltus , Alex F

Abstract

Entamoeba histolytica is an intestinal protozoan parasite and is the causative agent of amoebiasis. Upon entering the human host, cellular processes such as adhesion, phagocytosis, motility and secretion play a vital role in its propagation and pathogenicity. In other systems, each of these cellular processes is preceded by activation of signal transduction pathways, which often involve membrane phosphoinositides such as phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2). Little is known about phosphoinositide signaling in E. histolytica pathogenicity. In this study, we demonstrated that PI(4,5)P2 is localized to cholesterol-rich microdomains, lipid rafts, of the E. histolytica membrane and to the trailing edge of the polarized trophozoites. Since cholesterol correlates with virulence, we utilized cholesterol as a tool to determine the effect on subcellular and submembrane localization of PI(4,5)P2. Our data suggest that in E. histolytica, PI(4,5)P2 may signal from lipid rafts and cholesterol may play a role in triggering PI(4,5)P2-mediated signaling to enhance the motility of this pathogen.
We also utilized a functional genomics approach to identify genes that may operate in the phosphatidylinositol 3-kinase (PI3K) pathway in E. histolytica. Genes encoding a coactosin-like protein, EhCoactosin, and a serine-rich E. histolytica protein (SREHP) were identified in our screen. We utilized an independent genetic test to strengthen the connection between SREHP or EhCoactosin, and PI3K signaling. The assignment of these proteins as putative partners in PI3K signaling is novel. To our knowledge this is the first forward genetics screen adapted to reveal genes involved in the PI3K pathway in this pathogen.

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