Are protozoan metacaspases potential parasite killers?

Mechanisms concerning life or death decisions in protozoan parasites are still imperfectly understood. Comparison with higher eukaryotes has led to the hypothesis that caspase-like enzymes could be involved in death pathways. This hypothesis was reinforced by the description of caspase-related sequences in the genome of several parasites, including Plasmodium, Trypanosoma and Leishmania. Although several teams are working to decipher the exact role of metacaspases in protozoan parasites, partial, conflicting or negative results have been obtained with respect to the relationship between protozoan metacaspases and cell death. The aim of this paper is to review current knowledge of protozoan parasite metacaspases within a drug targeting perspective

Plasmodium falciparum metacaspase PfMCA-1 triggers a z-VAD-fmk inhibitable protease to promote cell death.

Activation of proteolytic cell death pathways may circumvent drug resistance in deadly protozoan parasites such as Plasmodium falciparum and Leishmania. To this end, it is important to define the cell death pathway(s) in parasites and thus characterize proteases such as metacaspases (MCA), which have been reported to induce cell death in plants and Leishmania parasites. We, therefore, investigated whether the cell death function of MCA is conserved in different protozoan parasite species such as Plasmodium falciparum and Leishmania major, focusing on the substrate specificity and functional role in cell survival as compared to Saccharomyces cerevisae. Our results show that, similarly to Leishmania, Plasmodium MCA exhibits a calcium-dependent, arginine-specific protease activity and its expression in yeast induced growth inhibition as well as an 82% increase in cell death under oxidative stress, a situation encountered by parasites during the host or when exposed to drugs such as artemisins. Furthermore, we show that MCA cell death pathways in both Plasmodium and Leishmania, involve a z-VAD-fmk inhibitable protease. Our data provide evidence that MCA from both Leishmania and Plasmodium falciparum is able to induce cell death in stress conditions, where it specifically activates a downstream enzyme as part of a cell death pathway. This enzymatic activity is also induced by the antimalarial drug chloroquine in erythrocytic stages of Plasmodium falciparum. Interestingly, we found that blocking parasite cell death influences their drug sensitivity, a result which could be used to create therapeutic strategies that by-pass drug resistance mechanisms by acting directly on the innate pathways of protozoan cell death

Place of the metacaspase pathway in Plasmodium falciparum apoptosis

Plasmodium falciparum est un protozoaire parasite responsable du paludisme causant la mort d’environ un million de personnes par an. La résistance médicamenteuse du parasite augmente la pathogénicité de cette maladie. Il est question ici d’explorer les mécanismes moléculaires impliqués dans la mort cellulaire programmée (apoptose) du parasite en présence de chloroquine (CQ) et de tester l’hypothèse qu’une résistance à la CQ peut s’expliquer en partie par une défaillance de ce mécanisme de mort. Dans un premier temps l’étude des marqueurs de l’apoptose (TUNEL, JC1, formes pyknotiques) montre qu’une souche sensible de parasite (3D7) à la CQ peut subir une apoptose en présence de CQ alors qu’une souche résistante (7G8) présente un défaut d’apoptose. Dans un deuxième temps nous montrons que la protéine PfMCA1 (P. falciparum métacaspase 1) présente une structure et une maturation protéolytique proche de celui des caspases faisant de cette protéine un candidat potentiellement impliqué dans l’apoptose du parasite. Dans un troisième temps nous montrons que l’expression du domaine catalytique de PfMCA1 dans la levure induit une mort cellulaire et un retard de croissance de la levure. Nous montrons également que PfMCA1 présente une activité enzymatique de type arginase alors que les effets induit par sa surexpression peuvent être inhibés par l’ajout d’un inhibiteur de protéases spécifiques des aspartates. Ces résultats suggèrent que PfMCA1 pourrait agir comme une protéine initiatrice induisant l’action d’une protéase effectrice spécifique des aspartates conduisant à la mort cellulaire. Cette hypothèse testée chez la levure reste à confirmée chez P. falciparumPlasmodium falciparum is a protozoan parasite responsible for malaria causing one million deaths per year. Drug resistance of the parasite increases the pathogenicity of this disease. In this thesis, it is question to explore the molecular pathway involved in programmed cell death (apoptosis) of the parasite in the presence of chloroquine (CQ) and to test the hypothesis that CQ resistance could be partly explained by a failure of such a mechanism. In a first step, we showed that a sensitive clone (3D7) exhibited the classical hallmarks of apoptosis (DNA fragmentation, mitochondrial depolarization) under a CQ pressure while a resistance clone failed to undergo apoptosis. In a second step we show that the protein PfMCA1 (P. falciparum metacaspase 1) has a structure and a processing similar to the well known caspases which are the key effectors of apoptosis for higher eukaryotic cells. In a third step we show that expression of the catalytic domain of PfMCA1 in yeast induces cell death and growth retardation of yeast. We show that PfMCA1 presented an arginine-specific protease activity while the effects induced by its overexpression were inhibited by an aspartate-specific protease inhibitor (z-VAD-fmk). These results suggest that PfMCA1 might act as an initiator protein inducing an aspartate-specific protease effector leading to cell death. This hypothesis tested in yeast remains to be confirmed in P. falciparu

L’innovation pédagogique dans les projets de l’Institut Carnot de l’Éducation Auvergne-Rhône-Alpes (ICÉ-AuRA)

International audienc

L’innovation pédagogique dans les projets de l’Institut Carnot de l’Éducation Auvergne-Rhône-Alpes (ICÉ-AuRA)

International audienc

L’innovation pédagogique dans les projets de l’Institut Carnot de l’Éducation Auvergne-Rhône-Alpes (ICÉ-AuRA)

International audienc