Cytosolic iron‐sulphur protein assembly is functionally conserved and essential in procyclic and bloodstream Trypanosoma brucei

Cytosolic and nuclear iron‐sulphur (Fe/S) proteins include essential components involved in protein translation, DNA synthesis and DNA repair. In yeast and human cells, assembly of their Fe/S cofactor is accomplished by the CIA (cytosolic iron‐sulphur protein assembly) machinery comprised of some 10 proteins. To investigate the extent of conservation of the CIA pathway, we examined its importance in the early‐branching eukaryote Trypanosoma brucei that encodes all known CIA factors. Upon RNAi‐mediated ablation of individual, early‐acting CIA proteins, no major defects were observed in both procyclic and bloodstream stages. In contrast, parallel depletion of two CIA components was lethal, and severely diminished cytosolic aconitase activity lending support for a direct role of the CIA proteins in cytosolic Fe/S protein biogenesis. In support of this conclusion, the T. brucei CIA proteins complemented the growth defects of their respective yeast CIA depletion mutants. Finally, the T. brucei CIA factor Tah18 was characterized as a flavoprotein, while its binding partner Dre2 functions as a Fe/S protein. Together, our results demonstrate the essential and conserved function of the CIA pathway in cytosolic Fe/S protein assembly in both developmental stages of this representative of supergroup Excavata.     

Light and dark in chromatin repair: repair of UV-induced DNA lesions by photolyase and nucleotide excision repair

Nucleotide excision repair (NER) and DNA repair by photolyase in the presence of light (photoreactivation) are the major pathways to remove UV-induced DNA lesions from the genome, thereby preventing mutagenesis and cell death. Photoreactivation was found in many prokaryotic and eukaryotic organisms, but not in mammals, while NER seems to be universally distributed. Since packaging of eukaryotic DNA in nucleosomes and higher order chromatin structures affects DNA structure and accessibility, damage formation and repair are coupled intimately to structural and dynamic properties of chromatin. Here, I review recent progress in the study of repair of chromatin and transcribed genes. Photoreactivation and NER are discussed as examples of how an individual enzyme and a complex repair pathway, respectively, access DNA lesions in chromatin and how these two repair processes fulfil complementary roles in removal of UV lesions. These repair pathways provide insight into the structural and dynamic properties of chromatin and suggest how other DNA repair processes could work in chromatin.     

Beyond immune escape: a variant surface glycoprotein causes suramin resistance in Trypanosoma brucei

Suramin is one of the first drugs developed in a medicinal chemistry program (Bayer, 1916), and it is still the treatment of choice for the hemolymphatic stage of African sleeping sickness caused by Trypanosoma brucei rhodesiense. Cellular uptake of suramin occurs by endocytosis, and reverse genetic studies with T. b. brucei have linked downregulation of the endocytic pathway to suramin resistance. Here we show that forward selection for suramin resistance in T. brucei spp. cultures is fast, highly reproducible and linked to antigenic variation. Bloodstream‐form trypanosomes are covered by a dense coat of variant surface glycoprotein (VSG), which protects them from their mammalian hosts' immune defenses. Each T. brucei genome contains over 2000 different VSG genes, but only one is expressed at a time. An expression switch to one particular VSG, termed VSG^Sur, correlated with suramin resistance. Reintroduction of the originally expressed VSG gene in resistant T. brucei restored suramin susceptibility. This is the first report of a link between antigenic variation and drug resistance in African trypanosomes.