HAT Complex Protein EAF6 - Role in Lifecycle Differentiation of Trypanosoma brucei

Abstract

During my project at the Schulz Lab, Harvey Mudd College, I investigated the role of the HAT complex protein EAF6 in the lifecycle differentiation of Trypanosoma brucei, the parasite responsible for African trypanosomiasis (“sleeping sickness”). This parasitic disease is known for its ability to evade the immune system through the expression of variant surface glycoproteins (VSG) in its bloodstream form, while the procyclic form expresses procyclin.

Our study focused on determining whether knocking down EAF6, a non-catalytic component of the histone acetyltransferase (HAT) complex, affects the parasite’s differentiation process. Using RNAi techniques, we designed and cloned vectors to knock down EAF6 expression in T. brucei. We then conducted differentiation experiments in both bloodstream and insect environments, monitored through EP1-GFP expression levels over three days using flow cytometry.

Results indicated no significant difference in differentiation between EAF6 knockdown (+Dox) and control (-Dox) conditions, although both showed slower differentiation compared to wild-type parasites. These findings suggest that while EAF6 plays a role in T. brucei lifecycle differentiation, our current RNAi clone may have had regulatory issues, necessitating further experimentation.

Future directions include repeating the experiments with independently transformed clones, performing Western blot analysis with EAF6 protein markers, and investigating the role of other HAT complex components like BDF6. This research contributes to our understanding of chromatin-interacting proteins in T. brucei and their potential as targets for therapeutic interventions against African trypanosomiasis.