An ADP-ribosyltransferase Alt of bacteriophage T4 negatively regulates the Escherichia coliMazF toxin of a toxin-antitoxin module

Prokaryotic toxin-antitoxin (TA) systems are linked to many roles in cell physiology, such as plasmid maintenance, stress response, persistence and protection from phage infection, and the activities of toxins are tightly regulated. Here, we describe a novel regulatory mechanism for a toxin of Escherichia coliTA systems. The MazF toxin of MazE-MazF, which is one of the best characterized type II TA systems, was modified immediately after infection with bacteriophage T4. Mass spectrometry demonstrated that the molecular weight of this modification was 542Da, corresponding to a mono-ADP-ribosylation. This modification disappeared in cells infected with T4 phage lacking Alt, which is one of three ADP-ribosyltransferases encoded by T4 phage and is injected together with phage DNA upon infection. In vivo and in vitro analyses confirmed that T4 Alt ADP-ribosylated MazF at an arginine residue at position 4. Finally, the ADP-ribosylation of MazF by Alt resulted in the reduction of MazF RNA cleavage activity in vitro, suggesting that it may function to inactivate MazF during T4 infection. This is the first example of the chemical modification of an E.coli toxin in TA systems to regulate activity. The MazF toxin of the MazE-MazF, one of the best-characterized type II TA systems, was modified immediately after infection with T4 phage, and a T4 ADP-ribosyltransferase Alt was responsible for this modification. Importantly, the ADP-ribosylation of MazF resulted in the reduction of its RNA cleavage activity, suggesting that it may function to inactivate MazF during T4 infection. This is the first example of the chemical modification of a toxin to regulate its activity.