Genomic and Proteomic Analysis of the Alkaline-Tolerance Response (AlTR) in Listeria monocytogenes 10403S

Information regarding the Alkaline Tolerance Response (AlTR) in Listeria monocytogenes is very limited. In this study, L. monocytogenes was shown to exhibit a significant adaptive alkaline tolerance response (AlTR) following a 1-h exposure to mild alkaline (pH 9.5), which is capable of protecting cells from subsequent lethal alkali stress (pH 12.0). Treatment of adapted cells with protein synthesis inhibitor chloramphenicol has revealed that AlTR is at least partially protein-dependent. In order to gain a more comprehensive perspective on the physiology and regulation of AlTR, we compared differential gene expression and protein content at pH 9.5 using microarray and two-dimensional (2D) gel electrophoresis, (combined with mass spectrometry) respectively. By interfacing the results of both approaches this study showed strong evidence that alkali tolerance response in L. monocytogenes functions as to minimize excess alkalisation and energy expenditures while mobilizing available carbon sources. Adaptive intracellular gene expression involved genes that are associated with virulence, the general stress response, cell division, and changes in cell wall structure and included many genes with unknown functions. The variability observed between results of cDNA arrays and 2D may account for posttranslational modifications. Interestingly, several alkali induced genes/proteins can provide a cross protective overlap to other types of stresses. Alkaline pH provides therefore L. monocytogenes with nonspecific multiple-stress resistance that may be vital for survival in the human gastrointestinal tract as well as within food processing systems where similar alkaline conditions as the ones used in this study prevail. Keywords: Transcriptomic and Proteomic Comparison Study Overnight grown Listeria monocytogenes 10403S culture (100 µl) was inoculated into a 250 ml Erlenmeyer flask with 100 ml of BHI, and cultures were incubated at 30° C with shaking at 200 rpm. Cells were harvested by centrifugation when the optical density at 600 nm reached ~0.4, and then rapidly transferred to prewarmed (30 ° C) alkali adjusted BHI pH 9.5 (Adapted cultures) or not adjusted BHI (Control cultures) for 60 min. The pH of BHI medium was adjusted to 9.5 with a glycine-NaOH-NaCl buffer. Control and alkali adapted cells were used for transcriptomic and proteomic studies. In array analysis several controls were employed to minimise technical and biological variations, and ensure the quality of the data, i.e. i) each ORF was present in duplicate in each array, ii) each RNA preparation was used to make probes for at least two-separate arrays with reversal of incorporated dye and iii) three independent RNA batches from each condition were analysed. Only differences of ≥ 2 fold change in the levels of gene expression, which also had SAM 'q' values of 0.05, were recorded as significant.