Proteomic Analysis Of Mycoplasma Pneumoniae Infected A549 Cell

Mycoplasma pneumoniae (MP) is a frequent cause of community-acquired respiratory infections in children and adults. Besides causing diseases in the respiratory system, MP has been implicated in several extra pulmonary complications arising from infection.Considerable progress has been made over the years in our understanding of MP pathogenesis. It has been shown that attachment of MP to the respiratory epithelium is a prerequisite for disease. Simultaneously, MP attachment induces the cells' inflammatory reaction and the host's immune response. Recently, some evidences have showed that MP can invade eukaryotic cells. Despite these advances in our understanding, the mechanisms underlying are still not fully clear. The rapid development of proteomic techniques has revolutionized our ability to study protein interactions and cellular changes on a global scale, allowing the discovery of previously unknown associations.To date, few studies have analyzed proteomics of MP infection. In this study, we investigated the effects of MP infection on A549 host cell protein profiles to elucidate the pathogenesis mechanism by using 2-dimensional gel electrophoresis (2-DE) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) technique. We used this proteomics approach in order to seek new pathogenesis-associated proteins.First, the A549 cell was infected by MP in different concentration. Effects of MP infection on A549 cell viability was determined by MTT test. It was found that MP infection had little effect on cell viability and cell proliferation below the concentration of 10 CFU/cell. Therefore, in the following experiments, unless otherwise specified, A549 cells were infected with MP at a 10 CFU/cell concentration. Second, high-resolution 2-DE was performed on the cellular proteins from A549 cells with or without MP-infection for 2h, 8h and 24h respectively. The gels were analyzed for quantitative spot comparisons with the image analysis software. Digitized images were analyzed by PDQuest software. Thirty-four protein spots were identified successfully. Compared to the control, 14 protein spots were up-regulated and 20 protein spots were down-regulated. The top biological functions of differential proteins were cytoskeleton followed by cell stress and cell DNA damage repair, signal transduction, transcriptional control, protein translation, modification and transportation, cellular metabolism, proliferation and differentiation. All of these proteins were involved in basal metabolism and biological process in cells. These findings suggested that the differential proteins identified in A549 cells with or without MP-infection might be implicated in MP pathogenesis.To verify the results of 2-D gel electrophoresis, LaminA/C and Ruvbl2 were chosen and subjected to Western blot analysis of cell lysate from M. pneumoniae-infected cells. The changes in protein abundance noted in the Western blot analysis were considerably consistent with the result obtained by proteomics, which demonstrates the accuracy of proteomic study.In conclusion, our data demonstrated that:MP infection may affect protein expression in A549 cells. Using proteomics approach to investigate differential proteome of A549 cells is a feasible strategy to seek pathogenesis of MP infection.