Trypanosoma brucei RAP1 Has Essential Functional Domains That Are Required for Different Protein Interactions
Marjia Afrin, Hanadi Kishmiri, Ranjodh Sandhu, M. A. G. Rabbani, Bibo Li
Abstract
Trypanosoma brucei causes human African trypanosomiasis and regularly switches its major surface antigen, VSG, to evade the host immune response. VSGs are expressed from subtelomeres in a monoallelic fashion. Tb RAP1, a telomere protein, is essential for cell viability and VSG monoallelic expression and suppresses VSG switching. Although Tb RAP1 has conserved functional domains in common with its orthologs from yeasts to mammals, the domain functions are unknown. RAP1 orthologs have pleiotropic functions, and interaction with different partners is an important means by which RAP1 executes its different roles. We have established a Cre-loxP-mediated conditional knockout system for Tb RAP1 and examined the roles of various functional domains in protein expression, nuclear localization, and protein-protein interactions. This system enables further studies of Tb RAP1 point mutation phenotypes. We have also determined functional domains of Tb RAP1 that are required for several different protein interactions, shedding light on the underlying mechanisms of Tb RAP1-mediated VSG silencing.