Suramin action in African trypanosomes involves a RuvB-like DNA helicase

Abstract

Suramin is one of the oldest drugs in use today. It is still the treatment of choice for the hemolymphatic stage of African sleeping sickness caused by Trypanosoma brucei rhodesiense and it is also used for surra in camels, caused by Trypanosoma evansi. Yet despite one hundred years of use, suramins mode of action is not fully understood. Suramin is a polypharmacologic molecule that inhibits diverse proteins. Here we demonstrate that a DNA helicase of the pontin/ruvB-like 1 family, termed T. brucei RuvBL1, is involved in suramin resistance in African trypanosomes. Bloodstream-form T. b. rhodesiense under long-term selection for suramin resistance acquired a homozygous point mutation, isoleucin-312 to valine, close to the ATP binding site of T. brucei RuvBL1. The introduction of this missense mutation, by reverse genetics, into drug-sensitive trypanosomes significantly decreased their sensitivity to suramin. Intriguingly, the corresponding residue of T. evansi RuvBL1 was found mutated in a suramin-resistant field isolate, in that case to a leucin. RuvBL1 (Tb927.4.1270) is predicted to build a heterohexameric complex with RuvBL2 (Tb927.4.2000). RNAi-mediated silencing of gene expression of either T. brucei RuvBL1 or RuvBL2 caused cell death within 72 h. At 36 h after induction of RNAi, bloodstream-form trypanosomes exhibited a cytokinesis defect resulting in the accumulation of cells with two nuclei and two or more kinetoplasts. Taken together, these data indicate that RuvBL1 DNA helicase is among the primary targets of suramin in African trypanosomes. Abstract ImportanceAfrican trypanosomes cause sleeping sickness in humans, nagana in cattle, and surra in camels - lethal diseases for which there is no vaccine and only few drugs. One of the drugs is suramin, developed by Bayer in 1916. Yet despite 100 years of use, suramins mode of action is not fully understood at the molecular level. Here we show that a DNA helicase is involved: Trypanosoma brucei rhodesiense (causative agent of sleeping sickness) selected for suramin resistance acquired a point mutation in the DNA helicase RuvBL1 that, when introduced to wild-type trypanosomes, reduced their sensitivity to suramin. Intriguingly, the same site in RuvBL1 was mutated also in a suramin-resistant field isolate of T. evansi (causative agent of surra). We further demonstrate that RuvBL1 is essential for proper cell division of T. brucei. Thus we conclude that inhibition of RuvBL1 contributes to the trypanocidal action of suramin.