Assembly and function of a bacterial genotoxin

Abstract

The tripartite cytolethal distending toxin (CDT) induces cell cycle arrest and apoptosis in eukaryotic cells. The subunits CdtA and CdtC associate with the nuclease CdtB to form a holotoxin that translocates CdtB into the host cell, where it acts as a genotoxin by creating DNA lesions. Here we show that the crystal structure of the holotoxin from Haemophilus ducreyi reveals that CDT consists of an enzyme of the DNase-I family, bound to two ricin-like lectin domains. CdtA, CdtB and CdtC form a ternary complex with three interdependent molecular interfaces, characterized by globular, as well as extensive non-globular, interactions. The lectin subunits form a deeply grooved, highly aromatic surface that we show to be critical for toxicity. The holotoxin possesses a steric block of the CdtB active site by means of a non-globular extension of the CdtC subunit, and we identify putative DNA binding residues in CdtB that are essential for toxin activity.

Cytolethal Distending Toxin Family Members Are Differentially Affected by Alterations in Host Glycans and Membrane Cholesterol

“…The identification of ricin-like lectin domains in CdtA and CdtC from structural and biochemical data first suggested that these subunits may interact with carbohydrates on the cell surface (13,26,27). Consistent with this hypothesis, CDT produced by Escherichia coli (Ec-CDT) was reported to require N-linked glycoproteins for binding and subsequent intoxication of HeLa cells (23).…”

“…CDT Cloning, Expression, and Purification-Cloning and expression of CDTs was based on the method previously described (13). Cultures of A. actinomycetemcomitans (Y4) and E. coli (S5) were obtained from ATCC and C. jejuni (81-176) from Patricia Guerry (Naval Medical Research Center, Silver Spring, MD).…”

Cytolethal Distending Toxin from Shiga Toxin-Producing Escherichia coli O157 Causes Irreversible G 2 /M Arrest, Inhibition of Proliferation, and Death of Human Endothelial Cells

“…The CdtA and CdtC polypeptides constitute the heterodimeric (B) subunit (15,35,36), which is required for CDT binding to target cells (15,36) and for the intracellular delivery of CdtB (15,35). This mechanism of action is supported by the crystal structure of CDT holotoxin from H. ducreyi (HdCDT) (38). The cellular response to CDT is associated with the accumulation of serine-20-phosphorylated (stabilized) tumor supressor protein p53, which is the key regulator of G 1 arrest (12,13,22), or of a tyrosine-15-and threonine-14-phosphorylated, inactive form of cyclin-dependent kinase cdc2, which accounts for G 2 arrest (10).…”

Cytolethal distending toxin A, B and C subunit proteins are necessary for the genotoxic effect of Escherichia coli CDT-V

“…The proposed role of CdtA and CdtC is the transport of CdtB to the target cell and the attachment of the toxin to the cell surface, and it has been verified in the case of Campylobacter jejuni (Lara-Tejero and Galán, 2001) and Aggregatibacter actinomycetemcomitans (Nesić et al, 2004;Yamada et al, 2006). However, there is increasing evidence suggesting that despite the similar genotoxic activity, there are differences between the molecular mechanisms used by the different CDT types in interaction with the host cell (McSweeney and Dreyfus, 2005;Eshraghi et al, 2010;Gargi et al, 2013).…”