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dc.contributor.advisorKappe, Stefan H
dc.contributor.authorAustin, Laura Stoffer
dc.date.accessioned2016-09-22T15:48:44Z
dc.date.available2016-09-22T15:48:44Z
dc.date.submitted2016-08
dc.identifier.otherAustin_washington_0250E_16322.pdf
dc.identifier.urihttp://hdl.handle.net/1773/37210
dc.descriptionThesis (Ph.D.)--University of Washington, 2016-08
dc.description.abstractMalaria is caused by eukaryotic parasites of the genus Plasmodium. They have coevolved with their mammalian hosts, significantly shaping both parasite and host genomes. It is the most deadly parasitic infection in the world, with approximately 200 million clinical episodes annually and nearly half a million deaths, mostly of small children. Disease is caused by parasite infection of host red blood cells; however, prior to this clinical phase, the parasite undergoes a clinically silent liver phase. While our understanding of this part of the Plasmodium life cycle has undergone tremendous development within the past two decades, much of the basic biology remains unknown. Previous work strongly suggests that the hepatocyte intracellular environment is critical for successful development of liver stage parasites and evasion of host defenses such as apoptosis of infected cells. Moreover, there is evidence to suggest that parasites are able to seek out certain hepatocytes that provide a more permissive environment for development. This work seeks to describe hepatocyte phenotypes that show increased susceptibility to Plasmodium liver stage infection, and to interrogate these hepatocytes to discover host molecular factors driving this differential susceptibility. We describe two phenotypes of differential susceptibility to Plasmodium in hepatocytes. First, Plasmodium preferentially infects and develops in polyploid hepatocytes. This increased susceptibility of highly polyploid hepatocytes is conserved for multiple Plasmodium species including the human malaria parasite P. falciparum. Susceptibility cannot be explained by differences in hepatocyte size or DNA replication. However, highly polyploid hepatocytes show increased density of surface proteins known to be important for parasite infection at the point of invasion. Secondly, phenotypic variability in infection exists between two closely related mouse substrains, BALB/cJ and BALB/cByJ. A consistent five-fold increase in liver stage burden is found in BALB/cByJ mice when compared to BALB/cJ. This difference is due to increased hepatocyte susceptibility to initial infection rather than immune system clearance or differential development in the liver and cannot be fully explained by previous known infection factors. A broad, unbiased search for host factors contributing to this susceptibility using RNA-Seq transcriptomics identified gene candidates including several known host membrane proteins for potential future work. Finally, interrogation of these phenotypes by reverse-phase microarray identified intracellular host factors, particularly the phosphorylated version of Ribosomal Protein S6 (RPS6) as being significantly and strongly upregulated in susceptible host hepatocytes. Blocking RPS6 phosphorylation by small molecule kinase inhibitors led to a drop in Plasmodium infection of hepatocytes. Additionally, mice with mutant RPS6 lacking phosphorylatable residues show lower average liver stage parasite burden than wild type littermates. Taken together, this body of work shows that Plasmodium parasites do not find hepatocytes to be equal. Hepatocytes with distinct molecular characteristics are clearly preferred as host cells. By inspecting the molecular differences found in hepatocytes of differential susceptibilities, we might identify new host factors important to malaria parasite liver infection and provide novel mechanistic insights into the processes of host cell selection and susceptibility to infection. Understanding the needs of the parasite during liver stage infection provides information useful for the development of new interventional strategies critical for the prevention and elimination of the early stages of malaria parasite infection.
dc.format.mimetypeapplication/pdf
dc.language.isoen_US
dc.subjectHepatocyte
dc.subjectLiver stage
dc.subjectMalaria
dc.subjectPlasmodium
dc.subjectPolyploidy
dc.subjectRibosomal Protein S6
dc.subject.otherParasitology
dc.subject.otherMolecular biology
dc.subject.otherCellular biology
dc.subject.otherpathobiology
dc.titleHepatocyte determinants of susceptibility to pre-erythrocytic Plasmodium infection
dc.typeThesis
dc.embargo.termsOpen Access


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