Comparative genomics of drug resistance in Trypanosoma brucei rhodesiense

Abstract

Trypanosoma brucei rhodesiense is one of the causative agents of human sleeping sickness, a fatal disease that is transmitted by tsetse flies and restricted to Sub-Saharan Africa. Here we investigate two independent lines of T. b. rhodesiense that have been selected with the drugs melarsoprol and pentamidine over the course of 2 years, until they exhibited stable cross-resistance to an unprecedented degree. We apply comparative genomics and transcriptomics to identify the underlying mutations. Only few mutations have become fixed during selection. Three genes were affected by mutations in both lines: the aminopurine transporter AT1, the aquaporin AQP2, and the RNA-binding protein UBP1. The melarsoprol-selected line carried a large deletion including the adenosine transporter gene AT1, whereas the pentamidine-selected line carried a heterozygous point mutation in AT1, G430R, which rendered the transporter non-functional. Both resistant lines had lost AQP2, and both lines carried the same point mutation, R131L, in the RNA-binding motif of UBP1. The finding that concomitant deletion of the known resistance genes AT1 and AQP2 in T. b. brucei failed to phenocopy the high levels of resistance of the T. b. rhodesiense mutants indicated a possible role of UBP1 in melarsoprol–pentamidine cross-resistance. However, homozygous in situ expression of UBP1-Leu131 in T. b. brucei did not affect the sensitivity to melarsoprol or pentamidine.Electronic supplementary materialThe online version of this article (doi:10.1007/s00018-016-2173-6) contains supplementary material, which is available to authorized users.

In vitro drug susceptibility of two strains of the wildlife trypanosome, Trypanosoma copemani : A comparison with Trypanosoma cruzi

“…Furthermore, T. copemani G1 has always been found in blood of marsupials while T. copemani G2 has also been found in tissues (Botero et al., 2013). Interestingly, we found that both strains of T. copemani exhibited significant differences in susceptibility to the different drugs used, supporting previous findings that genetic variation/gene varinats within species determines the degree of susceptibility to drugs (Campos et al., 2011, Plourde et al., 2012, Graf et al., 2013, Graf et al., 2016, Laffitte et al., 2016). Previous studies have shown an association between T. cruzi genetic diversity and their susceptibility to different drugs.…”

Exploiting the Achilles’ heel of membrane trafficking in trypanosomes

“…An unusual aminopurine transporter, TbAT1/P2, a member of the highly conserved Equilibrative Nucleoside Transporter family, facilitates uptake of both [37]. Deletion or mutation of the TbAT1/P2 gene reduces the sensitivity of T. brucei to diamidines, particularly diminazene, and melaminophenyl arsenicals [38, 39, 40, 41, 42]. …”

“…Further analyses revealed that aquaglyceroporin 2 ( AQP2 ) determines MPXR [44•, 45•]. Further, re-introduction of a wild-type TbAQP2 allele in even the most resistant strains fully restores drug sensitivity [45•, 46••, 47•, 41]. Indeed, expressing TbAQP2 in Leishmania mexicana promastigotes profoundly increases their sensitivity to both pentamidine and melarsoprol [45 • ].…”

ATrypanosoma bruceiORFeome-based Gain-of-Function Library reveals novel genes associated with melarsoprol resistance

“…Melarsoprol remains the only drug available for the treatment of second stage T. b. rhodesiense infection, yet it is highly toxic and difficult to administer (30). In addition, there have been increased reports of melarsoprol resistance and treatment failures in sub-Saharan Africa since the 1990s, only some of which have been explained by transporter mutations (34–37). Since melarsoprol has been widely studied, with both known and unknown genes associated with its cytotoxicity and treatment failures, it was selected for a T. brucei GoF library proof of principle genetic screen.…”