Two linked genes of Leishmania infantum encode tryparedoxins localised to cytosol and mitochondrion

Castro, Helena Carla; Sousa, Carla; Novais, Marta; Santos, Marta; Budde, Heike; Cordeiro-da-Silva, Anabela; Flohé, Leopold; Tomás, Ana M.

doi:10.1016/j.molbiopara.2004.02.015

Cited by 64 publications

(56 citation statements)

References 49 publications

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“…To prepare mTXNPx red , mTXNPx ox was incubated in the presence of 5 mM DTT or its physiological reducing system consisting of 200 μM NADPH, 0.5 U/mL L. infantum trypanothione reductase (TR), 50 μM trypanothione disulfide (TS 2 ; Bachem), and 2.5 μM L. infantum tryparedoxin 2 (TXN2) for 30 min at 30°C. TR and TXN2 were purified as previously described (37). Upon reduction, mTXNPx was diluted into the different assay buffers maintaining a DTT concentration between 0.2 and 1.5 mM in the final activity assay.…”

Section: Methodsmentioning

confidence: 99%

Mitochondrial peroxiredoxin functions as crucial chaperone reservoir inLeishmania infantum

Teixeira

Castro

Cruz

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Cytosolic eukaryotic 2-Cys-peroxiredoxins have been widely reported to act as dual-function proteins, either detoxifying reactive oxygen species or acting as chaperones to prevent protein aggregation. Several stimuli, including peroxide-mediated sulfinic acid formation at the active site cysteine, have been proposed to trigger the chaperone activity. However, the mechanism underlying this activation and the extent to which the chaperone function is crucial under physiological conditions in vivo remained unknown. Here we demonstrate that in the vector-borne protozoan parasite Leishmania infantum, mitochondrial peroxiredoxin (Prx) exerts intrinsic ATP-independent chaperone activity, protecting a wide variety of different proteins against heat stress-mediated unfolding in vitro and in vivo. Activation of the chaperone function appears to be induced by temperature-mediated restructuring of the reduced decamers, promoting binding of unfolding client proteins in the center of Prx's ringlike structure. Client proteins are maintained in a folding-competent conformation until restoration of nonstress conditions, upon which they are released and transferred to ATP-dependent chaperones for refolding. Interference with client binding impairs parasite infectivity, providing compelling evidence for the in vivo importance of Prx's chaperone function. Our results suggest that reduced Prx provides a mitochondrial chaperone reservoir, which allows L. infantum to deal successfully with protein unfolding conditions during the transition from insect to the mammalian hosts and to generate viable parasites capable of perpetuating infection.chaperone | Leishmania | peroxiredoxin

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Section: Methodsmentioning

confidence: 99%

Mitochondrial peroxiredoxin functions as crucial chaperone reservoir inLeishmania infantum

Teixeira

Castro

Cruz

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…Tryparedoxin peroxidase diminishes the inhibition of thiol redox metabolism by antimonial drugs by reactive oxygen species reduction (Wyllie et al 2004(Wyllie et al , 2008König and Fairlamb 2007;Mandal et al 2007), and is overexpressed in SSG-resistant clinical L. donovani (Wyllie et al 2010). The tryparedoxin locus itself may be associated with both host immunogenicity and drug resistance in Leishmania (Stober et al 2005), and in particular, with the murine B-cell response against L. infantum (Castro et al 2004;Cabral et al 2008). We discovered variation at three genes associated with redox metabolism: the tryparedoxin peroxidase gene (LdBPK_ 151140) showed signatures of selection on the L. donovani lineage (Supplemental Table S9) and also had a pattern of balanced variation in the 17 lines.…”

Section: )mentioning

confidence: 99%

Whole genome sequencing of multiple Leishmania donovani clinical isolates provides insights into population structure and mechanisms of drug resistance

Downing¹,

Imamura²,

Decuypere³

et al. 2011

Genome Res.

384

516

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Visceral leishmaniasis is a potentially fatal disease endemic to large parts of Asia and Africa, primarily caused by the protozoan parasite Leishmania donovani. Here, we report a high-quality reference genome sequence for a strain of L. donovani from Nepal, and use this sequence to study variation in a set of 16 related clinical lines, isolated from visceral leishmaniasis patients from the same region, which also differ in their response to in vitro drug susceptibility. We show that wholegenome sequence data reveals genetic structure within these lines not shown by multilocus typing, and suggests that drug resistance has emerged multiple times in this closely related set of lines. Sequence comparisons with other Leishmania species and analysis of single-nucleotide diversity within our sample showed evidence of selection acting in a range of surface-and transport-related genes, including genes associated with drug resistance. Against a background of relative genetic homogeneity, we found extensive variation in chromosome copy number between our lines. Other forms of structural variation were significantly associated with drug resistance, notably including gene dosage and the copy number of an experimentally verified circular episome present in all lines and described here for the first time. This study provides a basis for more powerful molecular profiling of visceral leishmaniasis, providing additional power to track the drug resistance and epidemiology of an important human pathogen.[Supplemental material is available for this article.]Leishmaniases are a complex of diseases that range from self-curing lesions to gross disfigurations and potentially deadly visceral disease. The diseases are caused by protozoan parasites that are transmitted by sandflies in 88 countries and infect an estimated 12 million people (www.who.int/leishmaniasis/en/). Parasites of the Leishmania genus are remarkably biologically, clinically, and epidemiologically diverse and present enormous differences in disease tropism. The mildest form is cutaneous leishmaniasis, which is caused by Leishmania major and other species, and is largely limited to lesions around the area of a sandfly bite-though a diffuse form can also occur. Disfiguring mucocutaneous leishmaniasis is due to the destruction of nasopharyngeal tissue by parasites such as L. braziliensis. More significantly, visceral leishmaniasis is caused by parasites of the L. donovani species complex that can spread to internal organs and cause death.In 2005, sequencing the genome of L. major identified 8311 protein-coding genes and provided a framework for future comparative genomic studies (Ivens et al. 2005). The genome elucidated the full structural architecture of Leishmania chromosomes, which includes an unusual pattern of genes distributed in large directional clusters. Subsequently, the genomes of L. braziliensis and L. infantum were described-the latter is a member of the L. donovani complex (Peacock et al. 2007). A detailed comparison of these first three Leishmania genomes re...

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“…Such differential lysis capacity of digitonin was used to increase the resolution and the depth of total cytosolic and intracellular proteome via sequential lysis of Leishmania parasites (Foucher et al 2006). In this study, we improved the method for the isolation of an enriched mitochondrial fraction previously described by Castro et al (2004). Purity fractions were estimated using luciferase activity as cytosolic marker and LimTXNPx as mitochondrial marker protein (Castro et al 2004;Foucher et al 2006).…”

Section: Introductionmentioning

confidence: 99%

“…In this study, we improved the method for the isolation of an enriched mitochondrial fraction previously described by Castro et al (2004). Purity fractions were estimated using luciferase activity as cytosolic marker and LimTXNPx as mitochondrial marker protein (Castro et al 2004;Foucher et al 2006). …”

Section: Introductionmentioning

confidence: 99%

Leishmania infantum: tuning digitonin fractionation for comparative proteomic of the mitochondrial protein content

Hide¹,

Ritleng²,

Brizard³

et al. 2008

Parasitol Res

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Leishmania infantum belongs to the Kinetoplastidae that is characterized by a specific mitochondrial DNA, the kinetoplast. This parasite is responsible for both benign cutaneous leishmaniasis and severe visceral leishmaniasis in humans. Molecular determinants of such differences in pathogenesis are not well understood, and the parasites as well as their hosts may contribute to the disease phenotype. Factors that help parasite to adapt its metabolism to nutritional conditions encountered in different location might play pivotal roles in controlling parasite development in these various host environments. Thus, we have decided to initiate studies aimed to compare the mitochondrial protein content of L. infantum. To avoid the drawback caused by the most abundant proteins such as tubulin and proteins of the cytoskeleton present in whole cell extract, we have decided to fractionate the subcellular components of the cells. Using both cytosolic and mitochondrial markers, we have improved a protein pre-fractionation protocol using digitonin that allowed us to generate an enriched mitochondrial fraction.

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Two linked genes of Leishmania infantum encode tryparedoxins localised to cytosol and mitochondrion

Cited by 64 publications

References 49 publications

Mitochondrial peroxiredoxin functions as crucial chaperone reservoir inLeishmania infantum

Mitochondrial peroxiredoxin functions as crucial chaperone reservoir inLeishmania infantum

Whole genome sequencing of multiple Leishmania donovani clinical isolates provides insights into population structure and mechanisms of drug resistance

Leishmania infantum: tuning digitonin fractionation for comparative proteomic of the mitochondrial protein content

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