Biogenesis of Leishmania major -Harboring Vacuoles in Murine Dendritic Cells

Abstract

In mammalian hosts, Leishmania sp. parasites are obligatory intracellular organisms that invade macrophages and dendritic cells (DC), where they reside in endocytic organelles termed parasitophorous vacuoles (PV). Most of the present knowledge of the characteristics of PV harboring Leishmania sp. is derived from studies with infected macrophages. Since DC play a key role in host resistance to leishmaniasis, there is a need to understand the properties and biogenesis of PV in Leishmania sp.-infected DC. Therefore, we determined the acquisition of endosomal and lysosomal molecules by Leishmania major-containing compartments in DC at different maturation stages, using fluorescence labeling and confocal microscopy. The results show that newly formed phagosomes in DC rapidly develop into late endosomal compartments. However, the small GTPase Rab7, which regulates late fusion processes, was found only in PV of mature bone marrow-derived DC (BMDC); it was absent in immature BMDC, suggesting an arrest of their PV biogenesis at the stage of late endosomes. Indeed, fusion assays with endocytic tracers demonstrated that the fusion activity of L. majorharboring PV toward lysosomes is higher in mature BMDC than in immature BMDC. The inhibition of PV-lysosome fusion in DC is dependent upon the viability and life cycle stage of the parasite, because live promastigotes blocked the fusion almost completely, whereas killed organisms and amastigotes induced a considerable level of fusion activity. The differences in the fusion competences of immature and mature DC may be relevant for their distinct functional activities in the uptake, transport, and presentation of parasite antigens.Protozoan parasites of the genus Leishmania are transmitted to mammalian hosts by phlebotomine sand flies. During the blood meal of an infected insect, the flagellated promastigote form gains access to host tissues and is rapidly phagocytosed, predominantly by macrophages in which the parasites transform into amastigotes devoid of the external flagellum. This process occurs within organelles termed parasitophorous vacuoles (PV). Amastigotes are well adapted to the conditions in the PV, allowing them to survive and multiply intracellularly (7).A critical part of the chain of events leading to degradation and death of intracellular microorganisms, including Leishmania sp. parasites, in macrophages is phagolysosome biogenesis. Newly formed phagosomes are unable to digest their contents and must therefore undergo a complex maturation process, which is driven by a series of sequential fusion events between phagosomes and early endosomes, late endosomes, and lysosomes (1, 10, 14). Each phagosome maturation stage is characterized by specific compartmental markers, such as the early endosomal markers CD71 and Rab4, the late endosomal markers Rab7 and CD68, and the late endosomal/lysosomal glycoproteins LAMP-1 and LAMP-2. These steps lead to the formation of the phagolysosome, an acidic compartment displaying the harsh and lytic environment needed to destroy mi...

Targeted extracellular signal-regulated kinase activation mediated by Leishmania amazonensis requires MP1 scaffold

“…2A, second panel) on the surface of this compartment. These data confirm previous findings indicating that PV maturation occurs within 1 hour of infection into a LAMP-2 positive, late endosomal/phagosomal compartment [18]. …”

“…In DC, newly-formed, Leishmania -containing phagosomes rapidly acquired late-endosomal markers including LAMP-1 and -2 [18]. During this time promastigotes transformed into amastigotes.…”

Cryptococcus neoformansEnters the Endolysosomal Pathway of Dendritic Cells and Is Killed by Lysosomal Components

“…For example, Mycobacterium tuberculosis can translocate from phagolysosomes to the cytosol of DCs, thus escaping killing and degradation (57). At certain stages of its life cycle, Leishmania major resides in DC endocytic compartments where it can escape lysosomal killing by blocking the fusion of lysosomes (24). Salmonella enterica serovar Typhimurium is found in DC compartments that lack lysosomal markers, suggesting that this organism pre- vents phagolysosomal fusion (15).…”

Leishmania hijacking of the macrophage intracellular compartments

“…In addition, increasing evidence indicates that neutrophils play a complex role in parasite recognition and are important regulators of the host anti‐ Leishmania immune responses . Leishmania promastigotes are capable of directly invading immature and mature DCs and inflammatory monocyte‐derived DCs , thereby residing within intracellular parasitophorous vacuoles. In immature DCs, the maturation of Leishmania ‐containing parasitophorous vacuoles showing acquisition of CD68 and lysosome‐associated membrane protein (LAMP)‐1 and ‐2 is arrested at the stage of the late endosome, leading to parasite survival .…”

“…Leishmania promastigotes are capable of directly invading immature and mature DCs and inflammatory monocyte‐derived DCs , thereby residing within intracellular parasitophorous vacuoles. In immature DCs, the maturation of Leishmania ‐containing parasitophorous vacuoles showing acquisition of CD68 and lysosome‐associated membrane protein (LAMP)‐1 and ‐2 is arrested at the stage of the late endosome, leading to parasite survival . In contrast, in mature DCs, the Leishmania ‐containing parasitophorous vacuole, acquiring in addition the small GTPase Rab7, fuses with lysosomes for parasite degradation .…”