Martin Taylor - Academia.edu (original) (raw)

Papers by Martin Taylor

Bioorganic Medicinal Chemistry Letters, Mar 1, 2012

A number of hydroxamic acid derivatives which inhibit human histone deacetylases were investigate... more A number of hydroxamic acid derivatives which inhibit human histone deacetylases were investigated for efficacy against cultured bloodstream form Trypanosoma brucei. Three out of the four classes tested displayed significant activity. The majority of compounds blocked parasite growth in the submicromolar range. The most potent was a member of the sulphonepiperazine series with an IC 50 of 34 nM. These results identify lead compounds with potential for the development of a novel class of trypanocidal agent.

Biochemical Journal, 2008

Background: There is little existing knowledge about actual quality of drugs provided by differen... more Background: There is little existing knowledge about actual quality of drugs provided by different providers in Nigeria and in many sub-Saharan African countries. Such information is important for improving malaria treatment that will help in the development and implementation of actions designed to improve the quality of treatment. The objective of the study was to determine the quality of drugs used for the treatment of malaria in a broad spectrum of public and private healthcare providers.

Molecular and Biochemical Parasitology, 1998

Antigenic variation in African trypanosomes continues to be one of the most elaborate and intrigu... more Antigenic variation in African trypanosomes continues to be one of the most elaborate and intriguing strategies ever devised by a protozoan parasite to avoid complete destruction by the immune defense of its mammalian host.

PLoS neglected tropical diseases, 2015

The neglected parasitic infection Chagas disease is rapidly becoming a globalised public health i... more The neglected parasitic infection Chagas disease is rapidly becoming a globalised public health issue due to migration. There are only two anti-parasitic drugs available to treat this disease, benznidazole and nifurtimox. Thus it is important to identify and validate new drug targets in Trypanosoma cruzi, the causative agent. T. cruzi expresses an ER-localised ascorbate-dependent peroxidase (TcAPx). This parasite-specific enzyme has attracted interest from the perspective of targeted chemotherapy. To assess the importance of TcAPx in protecting T. cruzi from oxidative stress and to determine if it is essential for virulence, we generated null mutants by targeted gene disruption. Loss of activity was associated with increased sensitivity to exogenous hydrogen peroxide, but had no effect on susceptibility to the front-line Chagas disease drug benznidazole. This suggests that increased oxidative stress in the ER does not play a significant role in its mechanism of action. Homozygous kn...

Background: Trypanosoma cruzi is a protozoan pathogen of major medical importance in Latin Americ... more Background: Trypanosoma cruzi is a protozoan pathogen of major medical importance in Latin America. It is also an early diverging eukaryote that displays many unusual biochemical features. The completion of the T. cruzi genome project has highlighted the need to extend the range of techniques available to study gene function. To this end we report the development of a stable tetracycline-dependent expression vector applicable to this parasite and describe in detail the parameters of the system.

Journal of biomolecular screening, 2015

The protozoan Trypanosoma cruzi is the causative agent of Chagas disease, one of the world's ... more The protozoan Trypanosoma cruzi is the causative agent of Chagas disease, one of the world's major neglected infections. Although development of improved antiparasitic drugs is considered a priority, there have been no significant treatment advances in the past 40 years. Factors that have limited progress include an incomplete understanding of pathogenesis, tissue tropism, and disease progression. In addition, in vivo models, which allow parasite burdens to be tracked throughout the chronic stage of infection, have been lacking. To address these issues, we have developed a highly sensitive in vivo imaging system based on bioluminescent T. cruzi, which express a red-shifted luciferase that emits light in the tissue-penetrating orange-red region of the spectrum. The exquisite sensitivity of this noninvasive murine model has been exploited to monitor parasite burden in real time throughout the chronic stage, has allowed the identification of the gastrointestinal tract as the major ...

International Journal for Parasitology, 2004

Erratum to "The amino terminal domain of a novel WD repeat protein from Trypanosoma cruzi contain... more Erratum to "The amino terminal domain of a novel WD repeat protein from Trypanosoma cruzi contains a non-canonical mitochondrial targeting signal" [Int. Fig. 5. Subcellular location of N-GFP in relation to an endoplasmic reticulum-specific marker. Panel 1, a metacyclic trypomastigote stained with TOTO-3 to identify DNA (blue). The intense staining identifies the kinetoplast. Panel 2, the same parasite visualised to identify the endoplasmic reticulum-localised protein BiP. Panel 3, localisation of N-GFP (green). Panel 4, merged images of panels 1-3. Panel 5, phase image. In panel 4, the yellow coloration at the posterior end of the cell indicates where the mitochondrion spirals over and under the endoplasmic reticulum. Bar, 5 mm. q doi of original article

species that it shares with the IPR suggests that no location contributes as much to the overall ... more species that it shares with the IPR suggests that no location contributes as much to the overall alpha diversity of the Indian and Pacific oceans as does the IPR.

Background: Trypanosoma cruzi is a protozoan pathogen of major medical importance in Latin Americ... more Background: Trypanosoma cruzi is a protozoan pathogen of major medical importance in Latin America. It is also an early diverging eukaryote that displays many unusual biochemical features. The completion of the T. cruzi genome project has highlighted the need to extend the range of techniques available to study gene function. To this end we report the development of a stable tetracycline-dependent expression vector applicable to this parasite and describe in detail the parameters of the system.

Human African trypanosomiasis (HAT) manifests in two stages of disease: firstly, haemolymphatic, ... more Human African trypanosomiasis (HAT) manifests in two stages of disease: firstly, haemolymphatic, and secondly, an encephalitic phase involving the central nervous system (CNS). New drugs to treat the second-stage disease are urgently needed, yet testing of novel drug candidates is a slow process because the established animal model relies on detecting parasitemia in the blood as late as 180 days after treatment. To expedite compound screening, we have modified the GVR35 strain of Trypanosoma brucei brucei to express luciferase, and have monitored parasite distribution in infected mice following treatment with trypanocidal compounds using serial, non-invasive, bioluminescence imaging. Parasites were detected in the brains of infected mice following treatment with diminazene, a drug which cures stage 1 but not stage 2 disease. Intravital multi-photon microscopy revealed that trypanosomes enter the brain meninges as early as day 5 postinfection but can be killed by diminazene, whereas those that cross the blood-brain barrier and enter the parenchyma by day 21 survived treatment and later caused bloodstream recrudescence. In contrast, all bioluminescent parasites were permanently eliminated by treatment with melarsoprol and DB829, compounds known to cure stage 2 disease. We show that this use of imaging reduces by two thirds the time taken to assess drug efficacy and provides a dual-modal imaging platform for monitoring trypanosome infection in different areas of the brain.

Bioorganic & Medicinal Chemistry, 2015

Dual submicromolar trypanocidal-antiplasmodial compounds have been identified by screening and ch... more Dual submicromolar trypanocidal-antiplasmodial compounds have been identified by screening and chemical synthesis of 4-aminoquinoline-based heterodimeric compounds of three different structural classes. In Trypanosoma brucei, inhibition of the enzyme trypanothione reductase seems to be involved in the potent trypanocidal activity of these heterodimers, although it is probably not the main biological target. Regarding antiplasmodial activity, the heterodimers seem to share the mode of action of the antimalarial drug chloroquine, which involves inhibition of the haem detoxification process. Interestingly, all of these heterodimers display good brain permeabilities, thereby being potentially useful for late stage human African trypanosomiasis. Future optimization of these compounds should focus mainly on decreasing cytotoxicity and acetylcholinesterase inhibitory activity.

Nucleic acids research, Jan 28, 2015

The metabolic network of a cell represents the catabolic and anabolic reactions that interconvert... more The metabolic network of a cell represents the catabolic and anabolic reactions that interconvert small molecules (metabolites) through the activity of enzymes, transporters and non-catalyzed chemical reactions. Our understanding of individual metabolic networks is increasing as we learn more about the enzymes that are active in particular cells under particular conditions and as technologies advance to allow detailed measurements of the cellular metabolome. Metabolic network databases are of increasing importance in allowing us to contextualise data sets emerging from transcriptomic, proteomic and metabolomic experiments. Here we present a dynamic database, TrypanoCyc (http://www.metexplore.fr/trypanocyc/), which describes the generic and condition-specific metabolic network of Trypanosoma brucei, a parasitic protozoan responsible for human and animal African trypanosomiasis. In addition to enabling navigation through the BioCyc-based TrypanoCyc interface, we have also implemented ...

Advances in Parasitology, 2011

It is almost 20 years since genetic manipulation of Trypanosoma cruzi was first reported. In this... more It is almost 20 years since genetic manipulation of Trypanosoma cruzi was first reported. In this time, there have been steady improvements in the available vector systems, and the applications of the technology have been extended into new areas. Episomal vectors have been modified to enhance the level of expression of transfected genes and to facilitate the sub-cellular location of their products. Integrative vectors have been adapted to allow the development of inducible expression systems and the construction of vectors which enable genome modification through telomere-associated chromosome fragmentation. The uses of reverse genetic approaches to dissect peroxide metabolism and the mechanisms of drug activity and resistance in T. cruzi are illustrated in this chapter as examples of how the technology has been used to investigate biological function. Although there remains scope to improve the flexibility of these systems, they have made valuable contributions towards exploiting the genome sequence data and providing a greater understanding of parasite biology and the mechanisms of infection.

Trends in Parasitology, 2014

Bioluminescence imaging is a non-invasive technique which can be used to monitor infections in re... more Bioluminescence imaging is a non-invasive technique which can be used to monitor infections in real-time. However, its utility is restricted by difficulties in detecting pathogens in deep tissue. 'Red-shifted' luciferases, which emit light of longer wavelength than standard bioluminescence-generating proteins, greatly enhance sensitivity, and have wide applicability for studying parasite infections.

Tetrahedron Letters, 2013

This article appeared in a journal published by Elsevier. The attached copy is furnished to the a... more This article appeared in a journal published by Elsevier. The attached copy is furnished to the author for internal non-commercial research and education use, including for instruction at the authors institution and sharing with colleagues.

Proceedings of the National Academy of Sciences, 2008

Nifurtimox and benznidazole are the front-line drugs used to treat Chagas disease, the most impor... more Nifurtimox and benznidazole are the front-line drugs used to treat Chagas disease, the most important parasitic infection in the Americas. These agents function as prodrugs and must be activated within the parasite to have trypanocidal effects. Despite >40 years of research, the mechanism(s) of action and resistance have remained elusive. Here, we report that in trypanosomes, both drugs are activated by a NADH-dependent, mitochondrially localized, bacterial-like, type I nitroreductase (NTR), and that down-regulation of this explains how resistance may emerge. Loss of a single copy of this gene in Trypanosoma cruzi, either through in vitro drug selection or by targeted gene deletion, is sufficient to cause significant cross-resistance to a wide range of nitroheterocyclic drugs. In Trypanosoma brucei, loss of a single NTR allele confers similar cross-resistance without affecting growth rate or the ability to establish an infection. This potential for drug resistance by a simple mechanism has important implications, because nifurtimox is currently undergoing phase III clinical trials against African trypanosomiasis.

Proceedings of the National Academy of Sciences, 2005

The capacity to synthesize vitamin C (ascorbate) is widespread in eukaryotes but is absent from h... more The capacity to synthesize vitamin C (ascorbate) is widespread in eukaryotes but is absent from humans. The last step in the biosynthetic pathway involves the conversion of an aldonolactone substrate to ascorbate, a reaction catalyzed by members of an FAD-dependent family of oxidoreductases. Here we demonstrate that both the African trypanosome, Trypanosoma brucei, and the American trypanosome, Trypanosoma cruzi, have the capacity to synthesize vitamin C and show that this reaction occurs in a unique single-membrane organelle, the glycosome. The corresponding T. brucei flavoprotein (TbALO) obeys Michaelis-Menten kinetics and can utilize both L-galactono-␥-lactone and D-arabinono-␥lactone as substrate, properties characteristic of plant and fungal enzymes. We could detect no activity toward the mammalian enzyme substrate L-gulono-␥-lactone. TbALO null mutants (bloodstream form) were found to display a transient growth defect, a trait that was enhanced when they were cultured in medium in which the essential serum component had been pretreated with ascorbate oxidase to deplete vitamin C. It is implicit, therefore, that bloodstream-form trypanosomes also possess a capacity for ascorbate transport.

PLoS Neglected Tropical Diseases, 2013

Background: Human African trypanosomiasis is caused by infection with parasites of the Trypanosom... more Background: Human African trypanosomiasis is caused by infection with parasites of the Trypanosoma brucei species complex, and threatens over 70 million people in sub-Saharan Africa. Development of new drugs is hampered by the limitations of current rodent models, particularly for stage II infections, which occur once parasites have accessed the CNS. Bioluminescence imaging of pathogens expressing firefly luciferase (emission maximum 562 nm) has been adopted in a number of in vivo models of disease to monitor dissemination, drug-treatment and the role of immune responses. However, lack of sensitivity in detecting deep tissue bioluminescence at wavelengths below 600 nm has restricted the wide-spread use of in vivo imaging to investigate infections with T. brucei and other trypanosomatids.

Nucleic Acids Research, 2011

Topoisomerase-II accumulates at centromeres during prometaphase, where it resolves the DNA catena... more Topoisomerase-II accumulates at centromeres during prometaphase, where it resolves the DNA catenations that represent the last link between sister chromatids. Previously, using approaches including etoposide-mediated topoisomerase-II cleavage, we mapped centromeric domains in trypanosomes, early branching eukaryotes in which chromosome segregation is poorly understood. Here, we show that in bloodstream form Trypanosoma brucei, RNAi-mediated depletion of topoisomerase-IIa, but not topoisomerase-IIb, results in the abolition of centromere-localized activity and is lethal. Both phenotypes can be rescued by expression of the corresponding enzyme from T. cruzi. Therefore, processes which govern centromere-specific topoisomerase-II accumulation/activation have been functionally conserved within trypanosomes, despite the long evolutionary separation of these species and differences in centromeric DNA organization. The variable carboxyl terminal region of topoisomerase-II has a major role in regulating biological function. We therefore generated T. brucei lines expressing T. cruzi topoisomerase-II truncated at the carboxyl terminus and examined activity at centromeres after the RNAi-mediated depletion of the endogenous enzyme. A region necessary for nuclear localization was delineated to six residues. In other organisms, sumoylation of topoisomerase-II has been shown to be necessary for regulated chromosome segregation. Evidence that we present here suggests that sumoylation of the T. brucei enzyme is not required for centromerespecific cleavage activity.

Bioorganic Medicinal Chemistry Letters, Mar 1, 2012

A number of hydroxamic acid derivatives which inhibit human histone deacetylases were investigate... more A number of hydroxamic acid derivatives which inhibit human histone deacetylases were investigated for efficacy against cultured bloodstream form Trypanosoma brucei. Three out of the four classes tested displayed significant activity. The majority of compounds blocked parasite growth in the submicromolar range. The most potent was a member of the sulphonepiperazine series with an IC 50 of 34 nM. These results identify lead compounds with potential for the development of a novel class of trypanocidal agent.

Biochemical Journal, 2008

Background: There is little existing knowledge about actual quality of drugs provided by differen... more Background: There is little existing knowledge about actual quality of drugs provided by different providers in Nigeria and in many sub-Saharan African countries. Such information is important for improving malaria treatment that will help in the development and implementation of actions designed to improve the quality of treatment. The objective of the study was to determine the quality of drugs used for the treatment of malaria in a broad spectrum of public and private healthcare providers.

Molecular and Biochemical Parasitology, 1998

Antigenic variation in African trypanosomes continues to be one of the most elaborate and intrigu... more Antigenic variation in African trypanosomes continues to be one of the most elaborate and intriguing strategies ever devised by a protozoan parasite to avoid complete destruction by the immune defense of its mammalian host.

PLoS neglected tropical diseases, 2015

The neglected parasitic infection Chagas disease is rapidly becoming a globalised public health i... more The neglected parasitic infection Chagas disease is rapidly becoming a globalised public health issue due to migration. There are only two anti-parasitic drugs available to treat this disease, benznidazole and nifurtimox. Thus it is important to identify and validate new drug targets in Trypanosoma cruzi, the causative agent. T. cruzi expresses an ER-localised ascorbate-dependent peroxidase (TcAPx). This parasite-specific enzyme has attracted interest from the perspective of targeted chemotherapy. To assess the importance of TcAPx in protecting T. cruzi from oxidative stress and to determine if it is essential for virulence, we generated null mutants by targeted gene disruption. Loss of activity was associated with increased sensitivity to exogenous hydrogen peroxide, but had no effect on susceptibility to the front-line Chagas disease drug benznidazole. This suggests that increased oxidative stress in the ER does not play a significant role in its mechanism of action. Homozygous kn...

Background: Trypanosoma cruzi is a protozoan pathogen of major medical importance in Latin Americ... more Background: Trypanosoma cruzi is a protozoan pathogen of major medical importance in Latin America. It is also an early diverging eukaryote that displays many unusual biochemical features. The completion of the T. cruzi genome project has highlighted the need to extend the range of techniques available to study gene function. To this end we report the development of a stable tetracycline-dependent expression vector applicable to this parasite and describe in detail the parameters of the system.

Journal of biomolecular screening, 2015

The protozoan Trypanosoma cruzi is the causative agent of Chagas disease, one of the world's ... more The protozoan Trypanosoma cruzi is the causative agent of Chagas disease, one of the world's major neglected infections. Although development of improved antiparasitic drugs is considered a priority, there have been no significant treatment advances in the past 40 years. Factors that have limited progress include an incomplete understanding of pathogenesis, tissue tropism, and disease progression. In addition, in vivo models, which allow parasite burdens to be tracked throughout the chronic stage of infection, have been lacking. To address these issues, we have developed a highly sensitive in vivo imaging system based on bioluminescent T. cruzi, which express a red-shifted luciferase that emits light in the tissue-penetrating orange-red region of the spectrum. The exquisite sensitivity of this noninvasive murine model has been exploited to monitor parasite burden in real time throughout the chronic stage, has allowed the identification of the gastrointestinal tract as the major ...

International Journal for Parasitology, 2004

Erratum to "The amino terminal domain of a novel WD repeat protein from Trypanosoma cruzi contain... more Erratum to "The amino terminal domain of a novel WD repeat protein from Trypanosoma cruzi contains a non-canonical mitochondrial targeting signal" [Int. Fig. 5. Subcellular location of N-GFP in relation to an endoplasmic reticulum-specific marker. Panel 1, a metacyclic trypomastigote stained with TOTO-3 to identify DNA (blue). The intense staining identifies the kinetoplast. Panel 2, the same parasite visualised to identify the endoplasmic reticulum-localised protein BiP. Panel 3, localisation of N-GFP (green). Panel 4, merged images of panels 1-3. Panel 5, phase image. In panel 4, the yellow coloration at the posterior end of the cell indicates where the mitochondrion spirals over and under the endoplasmic reticulum. Bar, 5 mm. q doi of original article

species that it shares with the IPR suggests that no location contributes as much to the overall ... more species that it shares with the IPR suggests that no location contributes as much to the overall alpha diversity of the Indian and Pacific oceans as does the IPR.

Background: Trypanosoma cruzi is a protozoan pathogen of major medical importance in Latin Americ... more Background: Trypanosoma cruzi is a protozoan pathogen of major medical importance in Latin America. It is also an early diverging eukaryote that displays many unusual biochemical features. The completion of the T. cruzi genome project has highlighted the need to extend the range of techniques available to study gene function. To this end we report the development of a stable tetracycline-dependent expression vector applicable to this parasite and describe in detail the parameters of the system.

Human African trypanosomiasis (HAT) manifests in two stages of disease: firstly, haemolymphatic, ... more Human African trypanosomiasis (HAT) manifests in two stages of disease: firstly, haemolymphatic, and secondly, an encephalitic phase involving the central nervous system (CNS). New drugs to treat the second-stage disease are urgently needed, yet testing of novel drug candidates is a slow process because the established animal model relies on detecting parasitemia in the blood as late as 180 days after treatment. To expedite compound screening, we have modified the GVR35 strain of Trypanosoma brucei brucei to express luciferase, and have monitored parasite distribution in infected mice following treatment with trypanocidal compounds using serial, non-invasive, bioluminescence imaging. Parasites were detected in the brains of infected mice following treatment with diminazene, a drug which cures stage 1 but not stage 2 disease. Intravital multi-photon microscopy revealed that trypanosomes enter the brain meninges as early as day 5 postinfection but can be killed by diminazene, whereas those that cross the blood-brain barrier and enter the parenchyma by day 21 survived treatment and later caused bloodstream recrudescence. In contrast, all bioluminescent parasites were permanently eliminated by treatment with melarsoprol and DB829, compounds known to cure stage 2 disease. We show that this use of imaging reduces by two thirds the time taken to assess drug efficacy and provides a dual-modal imaging platform for monitoring trypanosome infection in different areas of the brain.

Bioorganic & Medicinal Chemistry, 2015

Dual submicromolar trypanocidal-antiplasmodial compounds have been identified by screening and ch... more Dual submicromolar trypanocidal-antiplasmodial compounds have been identified by screening and chemical synthesis of 4-aminoquinoline-based heterodimeric compounds of three different structural classes. In Trypanosoma brucei, inhibition of the enzyme trypanothione reductase seems to be involved in the potent trypanocidal activity of these heterodimers, although it is probably not the main biological target. Regarding antiplasmodial activity, the heterodimers seem to share the mode of action of the antimalarial drug chloroquine, which involves inhibition of the haem detoxification process. Interestingly, all of these heterodimers display good brain permeabilities, thereby being potentially useful for late stage human African trypanosomiasis. Future optimization of these compounds should focus mainly on decreasing cytotoxicity and acetylcholinesterase inhibitory activity.

Nucleic acids research, Jan 28, 2015

The metabolic network of a cell represents the catabolic and anabolic reactions that interconvert... more The metabolic network of a cell represents the catabolic and anabolic reactions that interconvert small molecules (metabolites) through the activity of enzymes, transporters and non-catalyzed chemical reactions. Our understanding of individual metabolic networks is increasing as we learn more about the enzymes that are active in particular cells under particular conditions and as technologies advance to allow detailed measurements of the cellular metabolome. Metabolic network databases are of increasing importance in allowing us to contextualise data sets emerging from transcriptomic, proteomic and metabolomic experiments. Here we present a dynamic database, TrypanoCyc (http://www.metexplore.fr/trypanocyc/), which describes the generic and condition-specific metabolic network of Trypanosoma brucei, a parasitic protozoan responsible for human and animal African trypanosomiasis. In addition to enabling navigation through the BioCyc-based TrypanoCyc interface, we have also implemented ...

Advances in Parasitology, 2011

It is almost 20 years since genetic manipulation of Trypanosoma cruzi was first reported. In this... more It is almost 20 years since genetic manipulation of Trypanosoma cruzi was first reported. In this time, there have been steady improvements in the available vector systems, and the applications of the technology have been extended into new areas. Episomal vectors have been modified to enhance the level of expression of transfected genes and to facilitate the sub-cellular location of their products. Integrative vectors have been adapted to allow the development of inducible expression systems and the construction of vectors which enable genome modification through telomere-associated chromosome fragmentation. The uses of reverse genetic approaches to dissect peroxide metabolism and the mechanisms of drug activity and resistance in T. cruzi are illustrated in this chapter as examples of how the technology has been used to investigate biological function. Although there remains scope to improve the flexibility of these systems, they have made valuable contributions towards exploiting the genome sequence data and providing a greater understanding of parasite biology and the mechanisms of infection.

Trends in Parasitology, 2014

Bioluminescence imaging is a non-invasive technique which can be used to monitor infections in re... more Bioluminescence imaging is a non-invasive technique which can be used to monitor infections in real-time. However, its utility is restricted by difficulties in detecting pathogens in deep tissue. 'Red-shifted' luciferases, which emit light of longer wavelength than standard bioluminescence-generating proteins, greatly enhance sensitivity, and have wide applicability for studying parasite infections.

Tetrahedron Letters, 2013

This article appeared in a journal published by Elsevier. The attached copy is furnished to the a... more This article appeared in a journal published by Elsevier. The attached copy is furnished to the author for internal non-commercial research and education use, including for instruction at the authors institution and sharing with colleagues.

Proceedings of the National Academy of Sciences, 2008

Nifurtimox and benznidazole are the front-line drugs used to treat Chagas disease, the most impor... more Nifurtimox and benznidazole are the front-line drugs used to treat Chagas disease, the most important parasitic infection in the Americas. These agents function as prodrugs and must be activated within the parasite to have trypanocidal effects. Despite >40 years of research, the mechanism(s) of action and resistance have remained elusive. Here, we report that in trypanosomes, both drugs are activated by a NADH-dependent, mitochondrially localized, bacterial-like, type I nitroreductase (NTR), and that down-regulation of this explains how resistance may emerge. Loss of a single copy of this gene in Trypanosoma cruzi, either through in vitro drug selection or by targeted gene deletion, is sufficient to cause significant cross-resistance to a wide range of nitroheterocyclic drugs. In Trypanosoma brucei, loss of a single NTR allele confers similar cross-resistance without affecting growth rate or the ability to establish an infection. This potential for drug resistance by a simple mechanism has important implications, because nifurtimox is currently undergoing phase III clinical trials against African trypanosomiasis.

Proceedings of the National Academy of Sciences, 2005

The capacity to synthesize vitamin C (ascorbate) is widespread in eukaryotes but is absent from h... more The capacity to synthesize vitamin C (ascorbate) is widespread in eukaryotes but is absent from humans. The last step in the biosynthetic pathway involves the conversion of an aldonolactone substrate to ascorbate, a reaction catalyzed by members of an FAD-dependent family of oxidoreductases. Here we demonstrate that both the African trypanosome, Trypanosoma brucei, and the American trypanosome, Trypanosoma cruzi, have the capacity to synthesize vitamin C and show that this reaction occurs in a unique single-membrane organelle, the glycosome. The corresponding T. brucei flavoprotein (TbALO) obeys Michaelis-Menten kinetics and can utilize both L-galactono-␥-lactone and D-arabinono-␥lactone as substrate, properties characteristic of plant and fungal enzymes. We could detect no activity toward the mammalian enzyme substrate L-gulono-␥-lactone. TbALO null mutants (bloodstream form) were found to display a transient growth defect, a trait that was enhanced when they were cultured in medium in which the essential serum component had been pretreated with ascorbate oxidase to deplete vitamin C. It is implicit, therefore, that bloodstream-form trypanosomes also possess a capacity for ascorbate transport.

PLoS Neglected Tropical Diseases, 2013

Background: Human African trypanosomiasis is caused by infection with parasites of the Trypanosom... more Background: Human African trypanosomiasis is caused by infection with parasites of the Trypanosoma brucei species complex, and threatens over 70 million people in sub-Saharan Africa. Development of new drugs is hampered by the limitations of current rodent models, particularly for stage II infections, which occur once parasites have accessed the CNS. Bioluminescence imaging of pathogens expressing firefly luciferase (emission maximum 562 nm) has been adopted in a number of in vivo models of disease to monitor dissemination, drug-treatment and the role of immune responses. However, lack of sensitivity in detecting deep tissue bioluminescence at wavelengths below 600 nm has restricted the wide-spread use of in vivo imaging to investigate infections with T. brucei and other trypanosomatids.

Nucleic Acids Research, 2011

Topoisomerase-II accumulates at centromeres during prometaphase, where it resolves the DNA catena... more Topoisomerase-II accumulates at centromeres during prometaphase, where it resolves the DNA catenations that represent the last link between sister chromatids. Previously, using approaches including etoposide-mediated topoisomerase-II cleavage, we mapped centromeric domains in trypanosomes, early branching eukaryotes in which chromosome segregation is poorly understood. Here, we show that in bloodstream form Trypanosoma brucei, RNAi-mediated depletion of topoisomerase-IIa, but not topoisomerase-IIb, results in the abolition of centromere-localized activity and is lethal. Both phenotypes can be rescued by expression of the corresponding enzyme from T. cruzi. Therefore, processes which govern centromere-specific topoisomerase-II accumulation/activation have been functionally conserved within trypanosomes, despite the long evolutionary separation of these species and differences in centromeric DNA organization. The variable carboxyl terminal region of topoisomerase-II has a major role in regulating biological function. We therefore generated T. brucei lines expressing T. cruzi topoisomerase-II truncated at the carboxyl terminus and examined activity at centromeres after the RNAi-mediated depletion of the endogenous enzyme. A region necessary for nuclear localization was delineated to six residues. In other organisms, sumoylation of topoisomerase-II has been shown to be necessary for regulated chromosome segregation. Evidence that we present here suggests that sumoylation of the T. brucei enzyme is not required for centromerespecific cleavage activity.