The Role Of Protein Kinase YihE In The MazEF Mediated Bacterial Programmed Cell Death

Bacterial toxin-antitoxin(TA)modules are abundant genetic elements that encode a toxin protein capable of inhibiting cell growth and an antitoxin that counteracts the toxin.The MazEF TA system is one of the widely distributed type II TA systems,which mediates the programmed cell death(PCD)of bacterial communities under antibiotics and other stress conditions.Previous studies have shown that YihE kinase plays a protective role in the process of drug treatment by antagonizing the MazEF-mediated PCD.However,the detailed mechanism of YihE action is not yet clear.To further understand the function of YihE and the specific mode of action between YihE and the MazEF system,the following work was carried out in an attempt to clarify the specific pathways of YihE: 1)single mutants and double-mutants of YihE and factors related to the MazEF system were constructed,and the interaction between YihE and these factors was analyzed by measuring the susceptibility of these mutants to nalidixic acid(Nal,a kind of quinolones),which indicated that YihE did not interact directly with the MazEF system related factors;2)the difference of phosphorylated proteins between the wild-type strain and the yihE mutant after nalidixic acid treatment was examined,and it was found that the yihE mutation resulted in the disappearance of phosphorylation of four proteins and significant decrease in the phosphorylation levels of other two proteins,suggesting these proteins are potential targets of YihE;3)four potential target proteins of YihE were expressed and purified,and were subjected to in vitro phosphorylation.We find that YihE could phosphorylate the screened proteins in vitro.Based on these results,we proposed a model for YihE action: 1)YihE regulates the activity of Pgm and PpsA by phosphorylation,produce 6-p-Glucosamine and PEP to avoid cell death;2)phosphorylation of DnaK improves DnaK's ability to repair the misfolded protein caused by antibiotic pressure,and eventually avoid cell death.The study on the self-protection systems of bacteria to target their key factors(such as YihE)or related biological pathways is of great significance to develop adjuvant drugs to improve the bactericidal efficiency of antibiotics,which would provide a new way to improve the efficacy of current drugs.