Date of Completion

5-3-2017

Embargo Period

4-24-2017

Keywords

trypanosoma brucei, LC8, CITFA, RPB7, DYNLL1

Major Advisor

Arthur Gunzl

Associate Advisor

Stephen King

Associate Advisor

Bruce Mayer

Associate Advisor

Justin Radolf

Associate Advisor

Blanka Rogina

Field of Study

Biomedical Science

Degree

Doctor of Philosophy

Open Access

Open Access

Abstract

Trypanosoma brucei, a member of the early diverged phylogenetic order Kinetoplastida, is a vector-borne parasite that causes lethal disease in both humans and livestock. Unfortunately, progress has been slow on developing new treatments, and there is a need for new therapeutics, as current therapies have issues of resistance, toxicity, and difficult administration. In order to design new therapeutics, molecules and interactions unique to the parasite must be detailed, in hopes that some will afford suitable drug targets.

One unique process in T. brucei that might be targeted is RNA polymerase I-mediated transcription. T. brucei is unique in that RNA polymerase I not only transcribes ribosomal gene units, as in all other organisms, but is also used to transcribe gene arrays that encode its major cell surface proteins, namely the variant surface glycoprotein, or VSG in the mammalian bloodstream stage of the parasite. The importance of VSG to T. brucei is highlighted by the fact that interference with VSG mRNA rapidly halts bloodstream form culture growth and leads to the clearance of trypanosomes from infected mice. Thus, targeting proteins and interactions essential for VSG production is a valid strategy against T. brucei.

Chapter II details an investigation of the interaction between LC8 and a class I transcription factor A (CITFA) subunit, CITFA2, which was the focus of my thesis work. Both of these proteins, and their interaction, are essential for VSG transcription and trypanosome viability, and interrupting either protein or their interaction could be a potential anti-trypanosome therapy. Chapter III contains the generation of a method that allows for gene silencing using heterologous sequences, which was necessary for the work in chapter II. Chapter IV focuses on RPB7, an RNA pol II subunit, which was published to be utilized by RNA pol I for transcription. While this finding was intriguing, it contradicted our biochemical RNA pol I characterizations. Data presented in this chapter clearly demonstrated that RPB7 is not a subunit of RNA pol I and not required for the transcription process by this polymerase.

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