Project description:Mycoplasma genitalium is the causative agent of non-gonococcal, chlamydia-negative urethritis in men and has been linked to reproductive tract disease syndromes in women. As with other mycoplasmas, M. genitalium lacks many regulatory genes because of its streamlined genome and total dependence on a parasitic existence. Therefore, it is important to understand how gene regulation occurs in M. genitalium, particularly in response to environmental signals likely to be encountered in vivo. In this study, we developed an oligonucleotide-based microarray to investigate transcriptional changes in M. genitalium following osmotic shock. Using a physiologically relevant osmolarity condition (0.3 M sodium chloride), we identified 39 up-regulated and 72 down-regulated genes. Of the up-regulated genes, 21 were of unknown function and 15 encoded membrane-associated proteins. The majority of down-regulated genes encoded enzymes involved in energy metabolism and components of the protein translation process. These data provide insight into the in vivo response of M. genitalium to hyperosmolarity conditions and identify candidate genes that may contribute to mycoplasma survival in the urogenital tract.
Project description:Mycoplasma genitalium is the causative agent of non-gonococcal, chlamydia-negative urethritis in men and has been linked to reproductive tract disease syndromes in women. As with other mycoplasmas, M. genitalium lacks many regulatory genes because of its streamlined genome and total dependence on a parasitic existence. Therefore, it is important to understand how gene regulation occurs in M. genitalium, particularly in response to environmental signals likely to be encountered in vivo. In this study, we developed an oligonucleotide-based microarray to investigate transcriptional changes in M. genitalium following osmotic shock. Using a physiologically relevant osmolarity condition (0.3 M sodium chloride), we identified 39 up-regulated and 72 down-regulated genes. Of the up-regulated genes, 21 were of unknown function and 15 encoded membrane-associated proteins. The majority of down-regulated genes encoded enzymes involved in energy metabolism and components of the protein translation process. These data provide insight into the in vivo response of M. genitalium to hyperosmolarity conditions and identify candidate genes that may contribute to mycoplasma survival in the urogenital tract. To examine the effects of hyperosmolarity on M. genitalium transcription, four 50 ml cultures of strain G37 in 75 cm2 tissue culture flasks (Corning) were grown to exponential phase, as determined by medium colour change and colony density. Then, NaCl was added to three flasks to achieve final concentrations of 0.1, 0.2 and 0.3 M. Parallel cultures of M. genitalium in the absence of NaCl served as controls. All cultures were incubated for 1 h at 37M-bM-^DM-^C prior to RNA extraction. Experiments were repeated six times, which produced six independent RNA sample pairs from NaCl-treated cultures and control cultures for each NaCl condition. Dye swap was performed on three of six RNA pairs to minimize effects caused by biased labelling efficiencies.
Project description:Mycoplasma genitalium and M. pneumoniae are two significant mycoplasmas that infect the urogenital and respiratory tracts of humans. Despite distinct tissue tropisms, they both have similar pathogenic mechanisms and infect/invade epithelial cells in the respective regions and persist within these cells. However, the pathogenic mechanisms of these species in terms of bacterium-host interactions are poorly understood. To gain insights on this, we infected HeLa cells independently with M. genitalium and M. pneumoniae and assessed gene expression by whole transcriptome sequencing (RNA-seq) approach. The results revealed that HeLa cells respond to M. genitalium and M. pneumoniae differently by regulating various protein-coding genes. Though there is a significant overlap between the genes regulated by these species, many of the differentially expressed genes were specific to each species. KEGG pathway and signaling network analyses revealed that the genes specific to M. genitalium are more related to cellular processes. In contrast, the genes specific to M. pneumoniae infection are correlated with immune response and inflammation, possibly suggesting that M. pneumoniae has some inherent ability to modulate host immune pathways.