| During methylotrophic growth, single carbon compounds such as methanol are oxidized to formaldehyde en route to fulfilling the carbon and energy needs of the cell. Formaldehyde is a highly toxic compound that reacts readily with proteins and nucleic acids. Given the role of formaldehyde as a central intermediate in methylotrophic metabolism, it is hypothesized that methylotrophs have evolved specific mechanisms for handling formaldehyde stress. Viability assays were conducted to characterize the responses of the methylotrophic bacterium Methylobacterium extorquens AM1 to a variety of stressors across a range of concentrations. These results have defined stress conditions for microarray analysis to identify candidate genes involved in both formaldehyde-specific and general stress responses. Mutants were generated in ten genes whose transcripts were upregulated in microarray analysis under conditions of formaldehyde stress but not osmotic or oxidative stress. Of these, six mutants demonstrated increased sensitivity to formaldehyde stress relative to wild type. However, these six mutants were not altered in their ability to withstand osmotic, oxidative, or acetaldehyde stress. These results demonstrate that these genes, which include three putative transcriptional regulators and three genes of unknown function, are involved in protection from formaldehyde stress in the methylotroph M. extorquens AM1. Further microarray comparisons between these three putative transcriptional regulator mutants and wild type responses to formaldehyde stress suggest regulatory networks of genes involved in helping the cell respond to formaldehyde stress. |
