A bacterial cytotoxin identifies the RhoA Exchange Factor Net1 as a key effector in the response to DNA damage

Abstract

Background: Exposure of adherent cells to DNA damaging agents, such as the bacterial cytolethal distending toxin (COT) or ionizing radiations (IR), activates the small GTPase RhoA, which promotes the formation of actin stress fibers and delays cell death. The signalling intermediates that regulate RhoA activation and promote cell survival are unknown. Principal findings: We demonstrate that the nuclear RhOA-specific Guanine nucleotide Exchange Factor (GEF) Net1 becomes dephosphorylated at a critical inhibitory site in cells exposed to CDT or IR. Expression of a dominant negative Nat1 or Net1 knock down by iRNA prevented RhoA activation; inhibited the formation of stress fibers, and enhanced cell death, indicating that Net1 is required for this RhoA-mediated responses to genotoxic stress. The Net1 and. RhoA-dependent signals involved activation of the Mitogen-Activated Protein Kinase p38 and its downstream target MAPK-activated protein kinase 2. Significance: Our data highlight the importance of Net1 in controlling RhoA and p38 MAPK mediated cell survival in cells exposed to DNA damaginq agents and illustrate a molecular pathway whereby chronic exposure to a bacterial toxin may promote genomic instability. © 2008 Guerra et al.