Biological Functions And Mechanism Of Listeria Monocytogenes Lmo0734-mediated Oxidative Tolerance And Infection

Listeria monocytogenes(LM)is a Gram-positive facultative intracellular pathogen that can cause listeriosis.It is exquisitely well adapted to cope with various environmental changes including oxidative,osmotic,or acid stresses.A pivotal step in adapting to stresses in the various environment is the reprogramming of the transcriptional landscape to align gene expression with the physiological needs of Listeria,and this is achieved by a plurality of both protein and ribonucleic acid transcriptional regulators.In the previous studies on the transcriptome sequencing of Yjb H and Grx in Listeria,it was found that the gene cluster(lmo0735-lmo0739)with extremely significant differences in transcription level.Through the literature review,we know that lmo0734 is the transcriptional regulator of this gene cluster(lmo0735-lmo0739).Therefore,this study will focus on the biological functions and mechanism of Lmo0734-mediated oxidative tolerance and infection in host cells.1.Study on oxidative tolerance mediated by the transcriptional regulatory proteinLmo0734 of Listeria monocytogenesIn this study,we aimed to explain the molecular functions of the L.monocytogenes Lmo0734,to determine whether it contributes to biological processes related to bacterial anti-oxidative tolerance and infection.The results showed no significant change in the ability of proliferation and motility in the lmo0734 deletion of Listeria,and the colony morphology became serrated and not fruity.In the oxidative tolerance experiments,we found that ?lmo0734 cope with the resistance of metal ion oxidants(copper),and the sulfhydryl specific oxidant hydrazine(diamide)was enhanced.Therefore,we speculated that the oxidative tolerance of the environment and survival ability was enhanced in the lmo0734 deletion of Listeria monocytogenes.2.Study on the biological function of infection mediated by Listeria monocytogenes transcriptional regulatory protein Lmo0734According to the above experimental results,the oxidative tolerance of Listeria lmo0734 deletion strain was enhanced.We carried out experiments to infect different cell lines.The results showed that the intracellular growth of ?lmo0734 in macrophage RAW264.7 was significantly decreased at 2 h,5 h,and 8 h.The adhesion,invasion,and proliferation of ?lmo0734 in human intestinal epithelial cells(Caco-2)was decreased significantly,which was consistent with the results of the RAW264.7 macrophages.There was no significant change in the number of spots and the diameter of ?lmo0734 in mouse fibroblasts(L929).Finally,we used the mice infection model to analyze the pathogenicity of the Listeria Lmo0734 gene deletion strain.The results showed that the colonization of the Listeria Lmo0734 gene deletion strain was significantly decreased in the liver and spleen of mice for 24 and 48 h.In mice survival experiments,we found that the pathogenicity of ?lmo0734 was decreased by 60% compared to the wild-type strain(EGD-e).Furthermore,To study the regulatory relationship between Listeria monocytogenes transcriptional regulator lmo0734 and virulence factors,we analyzed the transcription level of Listeria virulence factor by real-time quantitative PCR.Western blotting was used to analyze the changes in virulence protein expression level of Listeria.The results showed that the transcription level of virulence factors decreased significantly in ?lmo0734.The protein expression levels of virulence factors Prf A and Plc A were significantly decreased.These results intuitively indicated that L.monocytogenes Lmo0734 is involved in bacterial oxidative tolerance and virulence regulationIn summary,this study showed for the first time that the Listeria Lmo0734 has negatively regulated oxidative tolerance and positively regulated virulence factors in ?lmo0734.The pathogenicity of mice was significantly reduced in ?lmo0734.Subsequently,the regulatory mechanism between Listeria Lmo0734 and virulence factors was deeply studied,and the transcription and expression levels of virulence island-1 in Listeria were significantly decreased.The results of this study help us to understand the biological functions and mechanisms of the Lmo0734-mediated anti-oxidative stress and infection of host cells.This study suggests a great significance for preventing and controlling foodborne intracellular pathogenic bacteria infection and ensuring human health and safety.