Biology subjects

Descoteaux, A.

Publications and source records attributed to Descoteaux, A..

7 recordsLinked to original sources

bioRxiv2026

Leishmania exploits the macrophage endoplasmic reticulum-shaping protein CLIMP-63 to modulate mitochondrial biogenesis and bioenergetics

In Leishmania-infected macrophages, interactions between parasitophorous vacuoles and the host cell endoplasmic reticulum are central to parasite replication. Whether the complex architecture of this organelle impacts these interactions remains however unknown. Here, we identify the macrophage endoplasmic reticulum-shaping protein CLIMP-63 as a host factor promoting Leishmania infection. Following parasite internalization, we observed a redistribution of CLIMP-63 to the vicinity of parasitophorous vacuoles in a process dependent on the virulence glycolipid lipophosphoglycan, and its dissociation from the mitochondrial network. Mechanistically, we found that CLIMP-63 is required for Leishmania-induced mitochondrial DNA replication, cristae biogenesis, and enhanced mitochondrial respiration, which are critical for the ability of the parasite to colonize macrophages. Collectively, our findings uncover a role for CLIMP-63 in supporting macrophage colonization by Leishmania and illustrate how this intracellular pathogen exploits the host cell endoplasmic reticulum to reprogram mitochondrial functions and bioenergetics. AUTHOR SUMMARYThe protozoan parasite Leishmania replicates in macrophages within parasitophorous vacuoles. These vacuoles establish interactions with various organelles and modulate their properties and functionality to adapt the host cell to their own needs. In this regard, Leishmania rewires host cell mitochondria to produce key metabolites and to modify their bioenergetic profile. We obtained evidence that Leishmania exploits the role of the macrophage endoplasmic reticulum architectural protein CLIMP-63 to stimulate mitochondrial biogenesis and respiration. The fact that CLIMP-63 plays a central role in the ability of Leishmania to colonize macrophage illustrates how this parasite manipulates the host cell endoplasmic reticulum to modulatet the function and properties of other organelles.

microbiology↗

bioRxiv2024

VAPA mediates lipid exchange between Leishmania amazonensis and host macrophages

Leishmania is a vacuolar pathogen that replicates within parasitophorous vacuoles inside host phagocytes. To promote its replication, Leishmania relies on a panoply of strategies to acquire macromolecules such as lipids from host macrophages. In this study, we have evaluated the role of VAPA, an endoplasmic reticulum-resident membrane protein involved in inter-organellar lipid transport, in macrophages infected with L. amazonensis. Following infection of bone marrow-derived macrophages with metacyclic L. amazonensis promastigotes, we observed that VAPA gradually associates with communal parasitophorous vacuoles. Knockdown of VAPA prevented the replication of L. amazonensis, which was accompanied by an impaired parasitophorous vacuole expansion. Using fluorescent ceramide, we established that VAPA is required for the transport of sphingolipids to the parasitophorous vacuoles and for its acquisition by L. amazonensis amastigotes. Proximity-ligation and immunoprecipitation assays revealed that L. amazonensis hijacks VAPA by disrupting its interactions with the lipid transfer proteins CERT and ORP1L. Finally, we found that VAPA is essential for the transfer of the Leishmania virulence glycolipid lipophosphoglycan from the parasitophorous vacuoles to the host cell endoplasmic reticulum. We propose that VAPA contributes to the ability of L. amazonensis to colonize macrophages by mediating bi-directional transfer of lipids essential for parasite replication and virulence between the parasitophorous vacuoles and the host cell endoplasmic reticulum. AUTHOR SUMMARYThe protozoan parasite Leishmania amazonensis replicates in macrophages, within communal parasitophorous vacuoles. To satisfy its various auxotrophies, this parasite must obtain macronutrients and metabolites from its host cell, including lipids. To salvage host sphingolipids, we obtained evidence that L. amazonensis exploits a macrophage nonvesicular lipid transport mechanism that requires the endoplasmic reticulum membrane protein VAPA. Moreover, we found that VAPA is also required for the transfer of the Leishmania virulence glycolipid lipophosphoglycan from the parasitophorous vacuole to the macrophage endoplasmic reticulum. The fact that VAPA is essential for L. amazonensis to colonize macrophages is consistent with the central role that VAPA plays in mediating bi-directional transfer of lipids and illustrates the importance of the host cell endoplasmic reticulum in this host-parasite interaction.

cell biology↗

bioRxiv2022

Revisiting Leishmania GP63 host cell targets reveals a limited spectrum of substrates

Colonization of host phagocytic cells by Leishmania metacyclic promastigotes involves several parasite effectors, including the zinc-dependent metalloprotease GP63. The major mode of action of this virulence factor entails the cleavage/degradation of host cell proteins. Given the potent proteolytic activity of GP63, identification of its substrates requires the adequate preparation of cell lysates to prevent artefactual degradation during cell processing. In the present study, we re-examined the cleavage/degradation of reported GP63 substrates when GP63 activity was efficiently neutralized during the preparation of cell lysates. To this end, we infected bone marrow-derived macrophages with either wild type, {Delta}gp63, and {Delta}gp63+GP63 L. major metacyclic promastigotes for various time points. We prepared cell lysates in the absence or presence of the zinc-metalloprotease inhibitor 1,10-phenanthroline and examined the levels and integrity of ten previously reported host cell GP63 substrates. Inhibition of GP63 activity with 1,10-phenanthroline during the processing of macrophages prevented the cleavage/degradation of several previously described GP63 targets, including PTP-PEST, mTOR, p65RelA, c-Jun, VAMP3, and NLRP3. Conversely, we confirmed that SHP-1, Synaptotagmin XI, VAMP8, and Syntaxin-5 are bona fide GP63 substrates. These results point to the importance of efficiently inhibiting GP63 activity during the preparation of Leishmania-infected host cell lysates. In addition, our results indicate that the role of GP63 in Leishmania pathogenesis must be re-evaluated. AUTHORS SUMMARYIn the protozoan parasite Leishmania, the abundant zinc-dependent metalloprotease GP63 is expressed at high levels at the surface of the promastigotes forms of the parasite. Upon phagocytosis by host macrophages, this metalloprotease is released from the parasites surface and spreads across the cytosol of infected cells. There, GP63 cleaves a number of host cell proteins involved in the control of host microbicidal function and in the regulation of immune responses, thereby contributing the ability of Leishmania to impair host defence mechanisms against infection. Given the abundance and powerful proteolytic activity of GP63, it is crucial to prevent artefactual proteolysis during processing of infected cells to identify genuine GP63 substrates. In this study, we found that inhibition of GP63 activity with 1,10-phenanthroline during the processing of macrophages prevented the degradation of several of previously identified GP63 substrates. These results uncover the importance of efficiently inhibiting GP63 activity during the preparation of Leishmania-infected host cell lysates.

microbiology↗

bioRxiv2021

Leishmania donovani metacyclic promastigotes impair phagosome properties in inflammatory monocytes

Leishmaniasis, a debilitating disease with clinical manifestations ranging from self-healing ulcers to life-threatening visceral pathologies, is caused by protozoan parasites of the Leishmania genus. These professional vacuolar pathogens are transmitted by infected sand flies to mammalian hosts as metacyclic promastigotes and are rapidly internalized by various phagocyte populations. Classical monocytes are among the first myeloid cells to migrate to infection sites. Recent evidence shows that recruitment of these cells contributes to parasite burden and to the establishment of chronic disease. However, the nature of Leishmania-inflammatory monocyte interactions during the early stages of host infection has not been well investigated. Here, we aimed to assess the impact of Leishmania donovani metacyclic promastigotes on antimicrobial responses within these cells. Our data showed that inflammatory monocytes were readily colonized by L. donovani metacyclic promastigotes, while infection with Escherichia coli was efficiently cleared. Upon internalization, metacyclic promastigotes inhibited superoxide production at the parasitophorous vacuole (PV) through a mechanism involving exclusion of NADPH oxidase subunits gp91phox and p47phox from the PV membrane. Moreover, we observed that unlike phagosomes enclosing zymosan particles, vacuoles containing parasites acidified poorly. Interestingly, whereas the parasite surface coat virulence glycolipid lipophosphoglycan was responsible for the inhibition of PV acidification, impairement of the NADPH oxidase assembly was independent of lipophosphoglycan and of the metalloprotease GP63. Collectively, these observations indicate that permissiveness of inflammatory monocytes to L. donovani may thus be related to the ability of this parasite to impair the microbicidal properties of phagosomes.

immunology↗

bioRxiv2020

Sec22b regulates inflammatory responses by controlling the nuclear translocation of NF-κB

Soluble NSF attachment receptor (SNARE) proteins regulate the vesicle transport machinery in phagocytic cells. Within the secretory pathway, Sec22b is an ER-Golgi intermediate compartment (ERGIC)-resident SNARE that controls phagosome maturation and function in macrophages and dendritic cells. The secretory pathway controls the release of cytokines and may also impact the secretion of nitric oxide (NO), which is synthesized by the Golgi-active inducible nitric oxide synthase (iNOS). Whether ERGIC SNARE Sec22b controls NO and cytokine secretion, is unknown. Using bone marrow-derived dendritic cells (BMDC), we demonstrated that iNOS colocalizes with ERGIC/Golgi markers, notably Sec22b and its partner syntaxin-5 (Stx5), in the cytoplasm and at the phagosome. Pharmacological blockade of the secretory pathway hindered NO and cytokine release, and inhibited NF-{kappa}B translocation to the nucleus. Importantly, RNAi-mediated silencing of Sec22b revealed that NO and cytokine production were abrogated at the protein and mRNA levels. This correlated with deregulated mitogen-activated protein kinase signalling and reduced nuclear translocation of NF-{kappa}B. We also found that Sec22b co-occurs with NF-{kappa}B in both the cytoplasm and nucleus, pointing to a role for this SNARE in the shuttling of NF-{kappa}B. Collectively, our data unveiled a novel function for the ER-Golgi, and its resident SNARE Sec22b, in the production and release of inflammatory mediators.

cell biology↗

bioRxiv2020

Binding of Leishmania infantum LPG to the midgut is not sufficient to define vector competence in Lutzomyia longipalpis sand flies

The major surface lipophosphoglycan (LPG) of Leishmania parasites is critical to vector competence in restrictive sand fly vectors by mediating Leishmania attachment to the midgut epithelium, considered essential to parasite survival and development. However, the relevance of LPG for sand flies that harbor multiple species of Leishmania remains elusive. We tested binding of Leishmania infantum wild type (WT), LPG-defective ({Delta}lpg1 mutants) and add-back lines ({Delta}lpg1 + LPG1) to sand fly midguts in vitro and their survival in Lutzomyia longipalpis sand flies in vivo. Le. infantum WT parasites attached to the Lu. longipalpis midgut in vitro with late-stage parasites binding to midguts in significantly higher numbers compared to early-stage stage promastigotes. {Delta}lpg1 mutants did not bind to Lu. longipalpis midguts, and this was rescued in the {Delta}lpg1 + LPG1 lines, indicating that midgut binding is mediated by LPG. When Lu. longipalpis sand flies were infected with either Le. infantum WT, {Delta}lpg1, or {Delta}lpg1 + LPG1 of the BH46 or BA262 strains, the BH46 {Delta}lpg1 mutant, but not the BA262 {Delta}lpg1 mutant, survived and grew to similar numbers as the WT and {Delta}lpg1 + LPG1 lines. Exposure of BH46 and BA262 {Delta}lpg1 mutants to blood engorged midgut extracts led to the mortality of the BA262 {Delta}lpg1 but not the BH46 {Delta}lpg1 parasites. These findings suggest that Le. infantum LPG protects parasites on a strain-specific basis early in infection, likely against toxic components of blood digestion, however, it is not necessary to prevent Le. infantum evacuation along with the feces in the permissive vector Lu. longipalpis. IMPORTANCEIt is well established that LPG is sufficient to define the vector competence of restrictive sand fly vectors to Leishmania parasites. However, the permissiveness of other sand flies to multiple Leishmania species suggests that other factors might define vector competence for these vectors. In this study, we investigated the underpinnings of Leishmania infantum survival and development in its natural vector Lutzomyia longipalpis. We found out that LPG-mediated midgut binding persists in late-stage parasites. This observation is paradigm-changing and suggests that only a subset of infective metacyclics lose their ability to attach to the midgut with implications for parasite transmission dynamics. However, our data also demonstrate that LPG is not a determining factor in Leishmania infantum retention in the midgut of Lutzomyia longipalpis, a permissive vector. Rather, LPG appears to be more important in protecting some parasite strains from the toxic environment generated during blood meal digestion in the insect gut. Thus, the relevance of LPG in parasite development in permissive vectors appears to be complex and should be investigated on a strain-specific basis.

microbiology↗

bioRxiv2019

Translational profiling of macrophages infected with Leishmania donovani identifies mTOR- and eIF4A-sensitive immune-related transcripts

The protozoan parasite Leishmania donovani (L. donovani) causes visceral leishmaniasis, a chronic infection which is fatal when untreated. While previous studies showed that L. donovani reprograms transcription to subvert host cell functions, it remains unclear whether the parasite also alters host mRNA translation to establish a successful infection. To assess this, we compared transcriptome-wide translation in primary mouse macrophages infected with L. donovani promastigotes or amastigotes using polysome-profiling. This identified ample selective changes in translation (3,127 transcripts) which were predicted to target central cellular functions by inducing synthesis of proteins related to chromatin remodeling and RNA metabolism while inhibiting those related to intracellular trafficking and antigen presentation. Parallel quantification of protein and mRNA levels for a set of transcripts whose translation was activated upon L. donovani infection (Papbpc1, Eif2ak2, and Tgfb) confirmed, as indicated by polysome-profiling, increased protein levels despite largely unaltered mRNA levels. Mechanistic in silico analyses suggested activated translation depending on the kinase mTOR (e.g. Pabpc1) and the RNA helicase eIF4A (e.g. Tgfb) during infection. Accordingly, treatment with mTOR inhibitors torin-1 or rapamycin reversed L. donovani-induced PABPC1 without affecting corresponding transcript levels. Similarly, the production of TGF-{beta} decreased in presence of the eIF4A inhibitor silvestrol despite unaltered Tgfb mRNA levels. Consistent with parasite modulation of host eIF4A-sensitive translation to promote infection, silvestrol suppressed L. donovani replication within macrophages. In contrast, parasite survival was favored under mTOR inhibition. In summary, infection-associated changes in translation of mTOR- and eIF4A-sensitive mRNAs contribute to modulate mRNA metabolism and immune responses in L. donovani-infected macrophages. Although the net outcome of such translation programs favours parasite propagation, individual translation programs appear to have opposing roles during L. donovani infection, thereby suggesting their selective targeting as key for therapeutic effects. Author SummaryFine-tuning the efficiency of mRNA translation into proteins allows cells to tailor their responses to stress without the need for synthesizing new mRNA molecules. It is well established that the protozoan parasite Leishmania donovani alters transcription of specific genes to subvert host cell functions. However, discrepancies between transcriptomic and proteomic data suggest that post-transcriptional regulatory mechanisms also contribute to modulate host gene expression programs during L. donovani infection. Herein, we report that one third of protein-coding mRNAs expressed in macrophages are differentially translated upon infection with L. donovani. Our computational analyses reveal that subsets of mRNAs encoding functionally related proteins share the same directionality of translational regulation, which is likely to impact metabolic and microbicidal activity of infected cells. We also show that upregulated translation of transcripts that encode central regulators of mRNA metabolism and inflammation is sensitive to the activation of mTOR or eIF4A during infection. Finally, we observe that inhibition of eIF4A activity reduces parasite survival within macrophages while selective blockade of mTOR has the opposite effect. Thus, our study points to a dual role for translational control of host gene expression during L. donovani infection and suggests that novel regulatory nodes could be targeted for therapeutic intervention.

immunology↗