Leishmania braziliensis: Strain-Specific Modulation of Phagosome Maturation.

Leishmania (Viannia) braziliensis is responsible for the largest number of American tegumentary leishmaniasis (ATL) in Brazil. ATL can present several clinical forms including typical (TL) and atypical (AL) cutaneous and mucocutaneous (ML) lesions. To identify parasite and host factors potentially associated with these diverse clinical manifestations, we first surveyed the expression of two virulence-associated glycoconjugates, lipophosphoglycan (LPG) and the metalloprotease GP63 by a panel of promastigotes of Leishmania braziliensis (L. braziliensis) strains isolated from patients with different clinical manifestations of ATL and from the sand fly vector. We observed a diversity of expression patterns for both LPG and GP63, which may be related to strain-specific polymorphisms. Interestingly, we noted that GP63 activity varies from strain to strain, including the ability to cleave host cell molecules. We next evaluated the ability of promastigotes from these L. braziliensis strains to modulate phagolysosome biogenesis in bone marrow-derived macrophages (BMM), by assessing phagosomal recruitment of the lysosome-associated membrane protein 1 (LAMP-1) and intraphagosomal acidification. Whereas, three out of six L. braziliensis strains impaired the phagosomal recruitment of LAMP-1, only the ML strain inhibited phagosome acidification to the same extent as the L. donovani strain that was used as a positive control. While decreased phagosomal recruitment of LAMP-1 correlated with higher LPG levels, decreased phagosomal acidification correlated with higher GP63 levels. Finally, we observed that the ability to infect and replicate within host cells did not fully correlate with the inhibition of phagosome maturation. Collectively, our results revealed a diversity of strain-specific phenotypes among L. braziliensis isolates, consistent with the high genetic diversity within Leishmania populations.

Acetyl-CoA Synthetase 2 Promotes Cell Migration and Invasion of Renal Cell Carcinoma by Upregulating Lysosomal-Associated Membrane Protein 1 Expression.

BACKGROUND/AIMS: Reprogramming energy metabolism is an emerging hallmark of many cancers, and this alteration is especially evident in renal cell carcinomas (RCCs). However, few studies have been conducted on lipid metabolism. This study investigated the function and mechanism of lipid metabolism-related acetyl-CoA synthetase 2 (ACSS2) in RCC development, cell migration and invasion. METHODS: Quantitative real-time PCR (qRT-PCR) was used to determine the expression of ACSS2 in cancer tissue and adjacent tissue. The inhibition of ACSS2 expression was achieved by RNA interference, which was confirmed by qRT-PCR and Western blotting. Cell proliferation and apoptosis were detected by a CCK8 assay and a flow cytometry analysis, respectively. Cell migration and invasion were determined by the scratch and transwell assays. Following the knockdown of ACSS2 expression, the expression of the autophagy-related factor LAMP1 was measured by qRT-PCR and Western blotting. RESULTS: Compared to adjacent tissues, ACSS2 expression was upregulated in RCC cancer tissues and positively correlated with metastasis. Inhibition of ACSS2 had no effect on RCC cell proliferation or apoptosis. However, decreased ACSS2 expression was found to inhibit RCC cell migration and invasion. ACSS2 was determined to promote the expression of LAMP1, which can also promote cell migration. This pathway may be considered a potential mechanism through which ACSS2 participates in RCC development. CONCLUSION: These data suggest that ACSS2 is an important factor for promoting RCC development and is essential for cell migration and invasion, which it promotes by increasing the expression of LAMP1. Taken together, these findings reveal a potential target for the diagnosis and treatment of RCC.

Molecular mechanisms of ATP secretion during immunogenic cell death.

The immunogenic demise of cancer cells can be induced by various chemotherapeutics, such as anthracyclines and oxaliplatin, and provokes an immune response against tumor-associated antigens. Thus, immunogenic cell death (ICD)-inducing antineoplastic agents stimulate a tumor-specific immune response that determines the long-term success of therapy. The release of ATP from dying cells constitutes one of the three major hallmarks of ICD and occurs independently of the two others, namely, the pre-apoptotic exposure of calreticulin on the cell surface and the postmortem release of high-mobility group box 1 (HMBG1) into the extracellular space. Pre-mortem autophagy is known to be required for the ICD-associated secretion of ATP, implying that autophagy-deficient cancer cells fail to elicit therapy-relevant immune responses in vivo. However, the precise molecular mechanisms whereby ATP is actively secreted in the course of ICD remain elusive. Using a combination of pharmacological screens, silencing experiments and techniques to monitor the subcellular localization of ATP, we show here that, in response to ICD inducers, ATP redistributes from lysosomes to autolysosomes and is secreted by a mechanism that requires the lysosomal protein LAMP1, which translocates to the plasma membrane in a strictly caspase-dependent manner. The secretion of ATP additionally involves the caspase-dependent activation of Rho-associated, coiled-coil containing protein kinase 1 (ROCK1)-mediated, myosin II-dependent cellular blebbing, as well as the opening of pannexin 1 (PANX1) channels, which is also triggered by caspases. Of note, although autophagy and LAMP1 fail to influence PANX1 channel opening, PANX1 is required for the ICD-associated translocation of LAMP1 to the plasma membrane. Altogether, these findings suggest that caspase- and PANX1-dependent lysosomal exocytosis has an essential role in ATP release as triggered by immunogenic chemotherapy.