Abstract
Autophagy is a bulk cell-degradation process that occurs through the lysosomal machinery, and many reports have shown that it participates in microbial pathogenicity. However, the role of autophagy in Clostridium difficile infection (CDI), the leading cause of antibiotics-associated diarrhea, pseudomembranous colitis and even death in severe cases, is not clear. Here we report that the major virulent factor toxin B (TcdB) of Clostridium difficile elicits a strong autophagy response in host cells through its glucosyltransferase activity. Using a variety of autophagy-deficient cell lines, i.e. HeLa/ATG7−/−, MEF/atg7−/−, MEF/tsc2−/−, we demonstrate that toxin-triggered autophagy inhibits host cell proliferation, which contributes to TcdB-caused cytopathic biological effects. We further show that both the PI3K complex and mTOR pathway play important roles in this autophagy induction process and consequent cytopathic event. Although the glucosyltransferase activity of TcdB is responsible for inducing both cell rounding and autophagy, there is no evidence suggesting the causal relationship between these two events. Taken together, our data demonstrate for the first time that the glucosyltransferase enzymatic activity of a pathogenic bacteria is responsible for host autophagy induction and the following cell growth arrest, providing a new paradigm for the role of autophagy in host defense mechanisms upon pathogenic infection.
Highlights
We found that TcdA, another key virulent factor of C. difficile[29,30,31], leads to autophagosome formation and induces autophagy in HeLa cells (Supplementary Fig. S1B,C)
Since the glucosyltransferase activity is responsible for both TcdB-triggered autophagy and cell-rounding, and it has been reported that extracellular matrix (ECM) detachment can induce autophagy[67], we investigated whether there is a causal relationship between autophagy induction and cell-rounding
We report for the first time that host autophagy can be triggered by C. difficile cytotoxin B through its glucosyltransferase activity
Summary
As a gram-positive anaerobe bacterium, C. difficile exerts its pathogenic effects mainly by producing two virulent factors, enterotoxin A (TcdA) and cytotoxin B (TcdB)[29,30,31] Both toxins enter cells via receptor-mediated endocytosis[32,33,34]. Their glucosyltransferase (GT) domains are subsequently released into the cytoplasm where they mono-glucosylate small GTPases of the Rho subfamily[35, 36], such as RhoA, Rac[1], Cdc[42], and TC10, by using the UDP-glucoses as co-substrates[37,38,39,40,41,42,43,44] These reactions lead to actin condensation and cell-rounding, membrane blebbing, and eventually cell death[45,46,47,48,49,50]. We provide evidence that host autophagy, triggered by TcdB from C. difficile through its glucosyltransferase activity, is critical for TcdB to inhibit host cell proliferation which plays as an important role in the biologic effects of TcdB55
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