Targeting TatD nuclease and the MAPK Pathway: Luteolin multifaceted approach against Mycoplasma gallisepticum infection

Nucleases are considered to be a key virulence factor of Mycoplasma gallisepticum (MG) and targeting nucleases is considered a novelly and potential direction for developing anti-MG drugs. The TatD nuclease localizes to the MG cell cytoplasm and can be released via extracellular vesicles (EVs), enabling it to participate in the immune evasion process. In this study, we explore the pathogenic mechanisms of TatD nuclease and screens for potential inhibitors, which could help overcome the limitations of MG treatment. In order to observe the function of TatD nuclease within cells, We utilized the transmembrane peptide Trans-activator (TAT) to construct the Trans-activator-TatD (TAT-TatD) recombinant protein. The effects of TAT-TatD recombinant protein were evaluated using DNA hydrolysis assays, Sytox staining and scanning electron microscopy. Utilizing network pharmacology and molecular docking techniques, the potential molecular mechanisms of luteolin (LUT) in targeting TatD nuclease and anti-MG were screened and explored. Results indicate that the TAT-TatD recombinant protein retains enzymatic activity and enters cells via the TAT domain, thereby inducing pyroptosis. This process is associated with the cGAS-STING pathway. Molecular docking and biomembrane layer interference experiments indicate that LUT is a potential inhibitor of the TatD nuclease. Not only that, LUT can also mitigate the cellular damage caused by TatD nuclease. It can further alleviate MG infection-induced tracheal inflammatory damage in chicks by regulating the MAPK signaling pathway. Take together, this study further investigates the pathogenic mechanism of TatD nuclease and provides strong support for developing anti-MG drugs based on nucleases to alleviate the limitations of MG treatment.