Anya Rushmer

The ability to sense both internal and external factors is crucial for bacteria to adapt to a changing environment. In the case of Methylorubrum extorquens PA1, an extensively studied facultative methylotroph, adapting to a toxic environment is just part of its metabolism. Methylotrophic bacteria produce formaldehyde (FA) as an intermediate of single carbon metabolism which can pose a threat to cell growth if concentrations get too high as FA is highly reactive and can destroy numerous important metabolites (e.g. proteins, DNA). Recent studies have shown that the M. extorquens genome encodes two important proteins, EfgA and TtmR, that are FA sensors and halt growth in the presence of FA. It is known that TtmR is a member of the MarR family of transcription factors and recent RNA sequencing data revealed that it alters expression of 75 genes, including EfgA. Additionally, experiments that have deleted TtmR from the M. extorquens genome and subjected this mutant to various growth experiments have uncovered a unique phenotype alluding to function of the protein. Most notably, when transitioning to a methylotrophic metabolism, ∆ttmR mutants experience an imbalance in FA production and a significant growth delay. Moreover, in contrast to wild-type M. extorquens who can only grow in FA concentrations up to 3mM, ∆ttmR mutants can grow in media containing up to 4mM FA. This study aims to understand the role of 3 different gene clusters identified in previous work (Mext_0563-66, Mext_2763-64, and Mext_3523) that are thought to be regulated by TtmR. In order to do this, markerless deletions were created via allelic exchange. Then, the mutants were subjected to both a transition to methylotrophy and a FA resistance growth experiment in comparison with wild-type and ∆ttmR to determine the differences in lag time, growth rate, and final OD.