Real‐time imaging of<i>Leishmania mexicana</i>‐infected early phagosomes: a study using primary macrophages generated from green fluorescent protein‐Rab5 transgenic mice

Lippuner, Christoph; Paape, Daniel; Paterou, Athina; Brand, Janko; Richardson, Melville; Smith, Andrew J.; Hoffmann, Kirstin; Brinkmann, Volker; Blackburn, C. Clare; Aebischer, Toni

doi:10.1096/fj.08-108712

Cited by 23 publications

(23 citation statements)

References 54 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…of infection, amastigotes, but not promastigotes, retained Rab5 staining. These results agree with studies in L. mexicana infection, in which amastigotes retain Rab5 longer than promastigotes, i.e., about one and two minutes, respectively [48]. The ability of parasites to hold on longer to Rab5 is LPG mediated, as LPG KO promastigotes retained Rab5 similar to amastigotes.…”

Section: Discussionsupporting

confidence: 90%

Stage-Specific Pathways of Leishmania infantum chagasi Entry and Phagosome Maturation in Macrophages

et al. 2011

View full text Add to dashboard Cite

The life stages of Leishmania spp. include the infectious promastigote and the replicative intracellular amastigote. Each stage is phagocytosed by macrophages during the parasite life cycle. We previously showed that caveolae, a subset of cholesterol-rich membrane lipid rafts, facilitate uptake and intracellular survival of virulent promastigotes by macrophages, at least in part, by delaying parasitophorous vacuole (PV)-lysosome fusion. We hypothesized that amastigotes and promastigotes would differ in their route of macrophage entry and mechanism of PV maturation. Indeed, transient disruption of macrophage lipid rafts decreased the entry of promastigotes, but not amastigotes, into macrophages (P<0.001). Promastigote-containing PVs were positive for caveolin-1, and co-localized transiently with EEA-1 and Rab5 at 5 minutes. Amastigote-generated PVs lacked caveolin-1 but retained Rab5 and EEA-1 for at least 30 minutes or 2 hours, respectively. Coinciding with their conversion into amastigotes, the number of promastigote PVs positive for LAMP-1 increased from 20% at 1 hour, to 46% by 24 hours, (P<0.001, Chi square). In contrast, more than 80% of amastigote-initiated PVs were LAMP-1+ at both 1 and 24 hours. Furthermore, lipid raft disruption increased LAMP-1 recruitment to promastigote, but not to amastigote-containing compartments. Overall, our data showed that promastigotes enter macrophages through cholesterol-rich domains like caveolae to delay fusion with lysosomes. In contrast, amastigotes enter through a non-caveolae pathway, and their PVs rapidly fuse with late endosomes but prolong their association with early endosome markers. These results suggest a model in which promastigotes and amastigotes use different mechanisms to enter macrophages, modulate the kinetics of phagosome maturation, and facilitate their intracellular survival.

show abstract

Section: Discussionsupporting

confidence: 90%

Stage-Specific Pathways of Leishmania infantum chagasi Entry and Phagosome Maturation in Macrophages

et al. 2011

View full text Add to dashboard Cite

show abstract

“…Taken together, these results unambiguously prove that Leishmania inhibits its transport to the lysosomes to survive in human macrophages. However, our results have also shown that Leishmania containing PV in mouse macrophages does not recruit and retain Rab5a after 24 h of infection whereas Leishmania containing PV in mouse macrophages is shown to recruit Rab5 in early time point of infection [52]. This is possibly due to the fact that Leishmania infection does not overexpress Rab5a in mouse macrophages.…”

Section: Discussionmentioning

confidence: 75%

Leishmania donovani resides in modified early endosomes by upregulating Rab5a expression via the downregulation of miR-494

2017

View full text Add to dashboard Cite

Several intracellular pathogens arrest the phagosome maturation in the host cells to avoid transport to lysosomes. In contrast, the Leishmania containing parasitophorous vacuole (PV) is shown to recruit lysosomal markers and thus Leishmania is postulated to be residing in the phagolysosomes in macrophages. Here, we report that Leishmania donovani specifically upregulates the expression of Rab5a by degrading c-Jun via their metalloprotease gp63 to downregulate the expression of miR-494 in THP-1 differentiated human macrophages. Our results also show that miR-494 negatively regulates the expression of Rab5a in cells. Subsequently, L. donovani recruits and retains Rab5a and EEA1 on PV to reside in early endosomes and inhibits transport to lysosomes in human macrophages. Similarly, we have also observed that Leishmania PV also recruits Rab5a by upregulating its expression in human PBMC differentiated macrophages. However, the parasite modulates the endosome by recruiting Lamp1 and inactive pro-CathepsinD on PV via the overexpression of Rab5a in infected cells. Furthermore, siRNA knockdown of Rab5a or overexpression of miR-494 in human macrophages significantly inhibits the survival of the parasites. These results provide the first mechanistic insights of parasite-mediated remodeling of endo-lysosomal trafficking to reside in a specialized early endocytic compartment.

show abstract

“…Fusion between PVs before 12 h of infection was seldom observed. Indeed, Lippuner and colleagues found that a minority of Rab5-positive L. mexicana PVs fuse with each other in the first hours of intracellular infection [30]. The fusion between L. amazonensis PVs did not contribute to sustain PV enlargement because fused PVs regained their dimensions, a process probably related to compensatory membrane recycling.…”

Section: Discussionmentioning

confidence: 99%

The Diverse and Dynamic Nature of Leishmania Parasitophorous Vacuoles Studied by Multidimensional Imaging

Real

Mortara

2012

PLoS Negl Trop Dis

View full text Add to dashboard Cite

An important area in the cell biology of intracellular parasitism is the customization of parasitophorous vacuoles (PVs) by prokaryotic or eukaryotic intracellular microorganisms. We were curious to compare PV biogenesis in primary mouse bone marrow-derived macrophages exposed to carefully prepared amastigotes of either Leishmania major or L. amazonensis. While tight-fitting PVs are housing one or two L. major amastigotes, giant PVs are housing many L. amazonensis amastigotes. In this study, using multidimensional imaging of live cells, we compare and characterize the PV biogenesis/remodeling of macrophages i) hosting amastigotes of either L. major or L. amazonensis and ii) loaded with Lysotracker, a lysosomotropic fluorescent probe. Three dynamic features of Leishmania amastigote-hosting PVs are documented: they range from i) entry of Lysotracker transients within tight-fitting, fission-prone L. major amastigote-housing PVs; ii) the decrease in the number of macrophage acidic vesicles during the L. major PV fission or L. amazonensis PV enlargement; to iii) the L. amazonensis PV remodeling after homotypic fusion. The high content information of multidimensional images allowed the updating of our understanding of the Leishmania species-specific differences in PV biogenesis/remodeling and could be useful for the study of other intracellular microorganisms.

show abstract

Real‐time imaging ofLeishmania mexicana‐infected early phagosomes: a study using primary macrophages generated from green fluorescent protein‐Rab5 transgenic mice

Cited by 23 publications

References 54 publications

Stage-Specific Pathways of Leishmania infantum chagasi Entry and Phagosome Maturation in Macrophages

Stage-Specific Pathways of Leishmania infantum chagasi Entry and Phagosome Maturation in Macrophages

Leishmania donovani resides in modified early endosomes by upregulating Rab5a expression via the downregulation of miR-494

The Diverse and Dynamic Nature of Leishmania Parasitophorous Vacuoles Studied by Multidimensional Imaging

Contact Info

Product

Resources

About