Abstract
In Helicobacter pylori infection, vacuolating cytotoxin (VacA)-induced mitochondrial damage leading to apoptosis is believed to be a major cause of cell death. It has also been proposed that VacA-induced autophagy serves as a host mechanism to limit toxin-induced cellular damage. Apoptosis and autophagy are two dynamic and opposing processes that must be balanced to regulate cell death and survival. Here we identify the low-density lipoprotein receptor-related protein-1 (LRP1) as the VacA receptor for toxin-induced autophagy in the gastric epithelial cell line AZ-521, and show that VacA internalization through binding to LRP1 regulates the autophagic process including generation of LC3-II from LC3-I, which is involved in formation of autophagosomes and autolysosomes. Knockdown of LRP1 and Atg5 inhibited generation of LC3-II as well as cleavage of PARP, a marker of apoptosis, in response to VacA, whereas caspase inhibitor, benzyloxycarbonyl-VAD-fluoromethylketone (Z-VAD-fmk), and necroptosis inhibitor, Necrostatin-1, did not inhibit VacA-induced autophagy, suggesting that VacA-induced autophagy via LRP1 binding precedes apoptosis. Other VacA receptors such as RPTPα, RPTPβ, and fibronectin did not affect VacA-induced autophagy or apoptosis. Therefore, we propose that the cell surface receptor, LRP1, mediates VacA-induced autophagy and apoptosis.
Highlights
Helicobacter pylori vacuolating cytotoxin (VacA) receptor(s) responsible for apoptotic cell death and autophagy has not been identified
We identify the low-density lipoprotein receptor-related protein-1 (LRP1) as the VacA receptor for toxin-induced autophagy in the gastric epithelial cell line AZ-521, and show that VacA internalization through binding to LRP1 regulates the autophagic process including generation of LC3-II from LC3-I, which is involved in formation of autophagosomes and autolysosomes
AZ-521 cells transfected with small-interfering RNAs (siRNAs) of LRP1 did not show significant toxin binding resulting in internalization, suggesting that LRP1 mediates VacA binding to the cell surface and facilitates its internalization
Summary
Helicobacter pylori VacA receptor(s) responsible for apoptotic cell death and autophagy has not been identified. Results: VacA-induced autophagy via low-density lipoprotein receptor-related protein-1 (LRP-1) binding precedes apoptosis. In Helicobacter pylori infection, vacuolating cytotoxin (VacA)-induced mitochondrial damage leading to apoptosis is believed to be a major cause of cell death. We identify the low-density lipoprotein receptor-related protein-1 (LRP1) as the VacA receptor for toxin-induced autophagy in the gastric epithelial cell line AZ-521, and show that VacA internalization through binding to LRP1 regulates the autophagic process including generation of LC3-II from LC3-I, which is involved in formation of autophagosomes and autolysosomes. VacA is internalized by endocytosis [6], which is inhibited by CagA [7, 8], and exerts multiple effects on susceptible cells, including vacuolation and mitochondrial damage, leading eventually to apoptosis (9 –13). Similar to RPTP␣ and RPTP, LRP1 mediates VacA internalization in AZ-521 cells, but in contrast to RPTP␣ and RPTP, LRP1 targeted downstream pathways leading to autophagy and apoptosis
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