LAMP-2 absence interferes with plasma membrane repair and decreases T. cruzi host cell invasion

Abstract

Trypanosoma cruzi enters host cells by subverting the mechanism of cell membrane repair. In this process, the parasite induces small injuries in the host cell membrane leading to calcium entry and lysosomal exocytosis, which are followed by compensatory endocytosis events that drive parasites into host cells. We have previously shown that absence of both LAMP-1 and 2, major components of lysosomal membranes, decreases invasion of T. cruzi into host cells, but the mechanism by which they interfere with parasite invasion has not been described. Here we investigated the role of these proteins in parasitophorous vacuole morphology, host cell lysosomal exocytosis, and membrane repair ability. First, we showed that cells lacking only LAMP-2 present the same invasion phenotype as LAMP1/2-/- cells, indicating that LAMP-2 is an important player during T. cruzi invasion process. Second, neither vacuole morphology nor lysosomal exocytosis was altered in LAMP-2 lacking cells (LAMP2-/- and LAMP1/2-/- cells). We then investigated the ability of LAMP-2 deficient cells to perform compensatory endocytosis upon lysosomal secretion, the mechanism by which cells repair their membrane and T. cruzi ultimately enters cells. We observed that these cells perform less endocytosis upon injury when compared to WT cells. This was a consequence of impaired cholesterol traffic in cells lacking LAMP-2 and its influence in the distribution of caveolin-1 at the cell plasma membrane, which is crucial for plasma membrane repair. The results presented here show the major role of LAMP-2 in caveolin traffic and membrane repair and consequently in T. cruzi invasion.

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

“…6H and 6I) when compared to wild type cells (Fig. 6G), similar to what was observed for T. cruzi infection with the same cell lines [25]. Additionally, our results indicate that Leishmania promastigotes are able to trigger calcium signaling in host cells from intracellular stores (Fig.…”

“…To better evaluate the role of lysosomes in cell infection and specifically address whether PM repair is important for cell invasion, we performed the infection of LAMP-2 knockout and LAMP-1/2 double knockout MEFs with LLa -RFP. These cells are known to be deficient in PM repair due to the accumulation of cholesterol and caveolin in lysosomes and, for this reason, are less susceptible to the invasion of T. cruzi [25]. The results (Fig.…”

“…Interestingly, nocodazole could not prevent infection by itself, as observed for T. cruzi invasion [21], which is probably due to the fact that mammalian cells already have a portion of their lysosomes pre-bound to the PM, which could be sufficient to allow parasite invasion [23]. Moreover, LAMP-2 and LAMP-1/2 knockout cells, which have modified lysosomes and impaired PM repair ability [25] were less susceptible to infection by L. amazonensis (Fig. 6H and 6I) when compared to wild type cells (Fig.…”

Leishmania amazonensis hijacks host cell lysosomes involved in plasma membrane repair to induce invasion in fibroblasts

“…To better evaluate the role of lysosomes in cell infection and specifically address whether PM repair is important for cell invasion, we performed the LLa-RFP infection in LAMP2-knockout and LAMP1/2 double-knockout MEFs. These cells are known to be deficient in PM repair due to the accumulation of cholesterol and caveolin in lysosomes and, for this reason, are less susceptible to the invasion of T. cruzi (Couto et al, 2017). The results ( Fig.…”

How Does the Main Infective Stage ofT. cruziEnter and Avoid Degradation in Host Cells? A Description of the Pathways and Organelles Involved on These Processes

“…This decrease in entry is not accompanied by inhibition of lysosome exocytosis to the repair region as well as phenotypic modification of the host cell or generated PV. It is believed that the decrease in parasite entry is due to problems in caveolin-mediated endocytosis and in calcium efflux [67]. Considering that there are two distinct pools of lysosomes (cortical and internal) in mammalian cells, Hissa and Andrade used cardiomyocytes and observed, through the use of specific inhibitors, that trypomastigotes enter into this cell type mainly recruiting lysosomes from the more internal cell layer (perinuclear) [52].…”