Queiroz-Oliveira, T. | Search bioRxiv

bioRxiv2025

Leishmania amazonensis amastigotes invade non-phagocytic cells via highly localized parasite-induced actin remodeling.

Intracellular parasites are pathogens that must invade and persist within host cells. In the case of Leishmania spp., it is generally assumed that the parasite must be phagocytosed to further establish residence within professional phagocytic cells. However, several studies report the presence of Leishmania spp. in non-phagocytic cells, highlighting their ability to enter cells independently of classical phagocytosis. Indeed, we have recently demonstrated that promastigotes, the infective form transmitted to hosts during the bite of the insect vector, subvert a lysosome-exocytosis-dependent membrane repair pathway to invade fibroblasts. Here, we investigate the invasion of non-phagocytic cells by L. amazonensis amastigotes, the infective form directly responsible for host-to-vector transmission, disease generation and parasite dissemination during infection in humans and other mammalian hosts. Our results show that amastigotes rapidly induce their entry into cells lacking classical phagocytic capability, where they survive, multiply, and persist. Invasion depends on parasite-induced actin remodeling confined to the parasite-host plasma membrane contact site, with localized recruitment of Rho GTPases that fuel actin polymerization at invasion foci. Our findings highlight the remarkable ability of Leishmania amastigotes to induce their own internalization into virtually any cell type, bypassing the need for conventional phagocytosis. This property may profoundly influence parasite biology by enabling amastigotes to cross cellular barriers, disseminate, and silently establish infection in diverse host cells. Importantly, when considering non-phagocytic cells, our results demonstrate that Leishmania amazonensis employs distinct, stage-specific invasion mechanisms: promastigotes co-opt host cell lysosomes, whereas amastigotes depend on F-actin dynamics. SUMMARYHere we elucidate key aspects of the mechanism by which Leishmania amazonensis amastigotes invade non-phagocytic cells.

microbiology↗

bioRxiv2018

Unveiling Leishmania invasion of fibroblasts: calcium signaling, lysosome recruitment and exocytosis culminate with actin-independent invasion.

Intracellular parasites of the genus Leishmania are the causative agents of human leishmaniasis, a widespread emergent tropical disease. The parasite is transmitted by the bite of a hematophagous sandfly vector that inoculates motile flagellated promastigote forms into the dermis of the mammalian host. After inoculation, parasites are ultimately captured by macrophages and multiply as round-shaped amastigote forms. Macrophages seem not to be the first infected cells since parasites were observed invading neutrophils first whose leishmania-containing apoptotic bodies were latter captured by macrophages, thereby becoming infected. The fact that Leishmania spp are able to live and replicate inside immune phagocytic cells and that macrophages are the main cell type found infected in chronicity created the perception that Leishmania spp are passive players waiting to be captured by phagocytes. However, several groups have described the infection of non-phagocytic cells in vivo and in vitro. The objective of this work was to study the cellular mechanisms involved in the invasion of non-professional phagocytes by Leishmania. We show that promastigotes of L.amazonensis actively induces invasion in fibroblasts without cytoskeleton activity, thus by a mechanism that is distinct from phagocytosis. Inside fibroblasts parasites transformed in amastigotes, remained viable for at least two weeks and re-transformed in promastigotes when returned to insect vector conditions. Similarly to what was observed for T. cruzi, infection involves calcium signaling, recruitment and exocytosis of lysosomes involved in plasma membrane repair and lysosome-triggered endocytosis. Conditions that alter lysosomal function such as cytochalasin-D and brefeldin-A treatment or the knockout of host cell lysosomal proteins LAMP-1 and 2 dramatically affected invasion. Likewise, triggering of lysosomal exocytosis and lysosome-dependent plasma membrane repair by low doses of streptolysin-O dramatically increased parasite entry. Together our results show that L.amazonensis promastigotes are able to take advantage of calcium-dependent lysosomal exocytosis and lysosome-induced endocytosis to invade and persist in non-phagocytic cells.\n\nAUTHOR SUMMARYIntracellular parasites of the genus Leishmania are the causative agents of leishmaniasis. The disease is transmitted by the bite of a sand fly vector which inoculates the parasite into the skin of mammalian hosts, including humans. During chronic infection the parasite lives and replicates inside phagocytic cells, notably the macrophages. An interesting but overlooked finding on Leishmania infection is that non-phagocytic cells have also been found infected by amastigotes. Nevertheless, the mechanisms by which Leishmania invades non-phagocytic cells were not studied to date. Here we show that L. amazonensis can actively induce their own entry into fibroblasts independently of actin cytoskeleton activity, thus by a mechanism that is distinct from phagocytosis. Invasion involves subversion of host cell functions such as calcium signaling and recruitment and exocytosis of host cell lysosomes involved in plasma membrane repair and whose positioning and content interfere in invasion. Parasites were able to replicate and remained viable in fibroblasts, suggesting that cell invasion trough the mechanism demonstrated here could serve as a parasite hideout and reservoir, facilitating infection amplification and persistence.

cell biology↗