Discussion

Trypanosoma brucei relies on a dense Variant Surface Glycoprotein (VSG) coat for survival in the host bloodstream.  High VSG expression within an expression site body (ESB) is mediated by RNA polymerase I (Pol I), which normally exclusively transcribes rDNA.  Pol I inhibitors are currently in clinical trials against cancer, as they target cells that are reliant upon high levels of Pol I transcription.  As T. brucei also relies on Pol I transcription of a single VSG gene for survival within the host, we investigated Pol I inhibitors quarfloxin, CX-5461, and BMH-21 for selective efficacy.  Cytotoxicity assays showed all three Pol I inhibitors have IC50 concentrations in the nanomolar range in T. brucei, and selective toxicity compared with mammalian cells.  Parasite growth inhibition was due to rapid and specific inhibition of Pol I transcription, as rRNA precursor transcript was reduced to 2-3% and VSG precursor transcript to 6-9% normal levels within 15 minutes incubation with these substances.  Incubation with Pol I inhibitors also resulted in rapid loss of the ESB and disintegration of the nucleolus within one hour.  As ESB loss followed the rapid reduction in Pol I precursor transcripts, this suggests the ESB is a ‘transcription-nucleated’ subnuclear structure.  In addition to providing insight into Pol I transcription and ES control, Pol I inhibitors potentially also provide new leads against trypanosomiasis.