AbstractMethanotrophs oxidize most of the methane (CH₄) produced in natural and anthropogenic ecosystems. Often living close to soil surfaces, these microorganisms must frequently adjust to temperature change. While many environmental studies have addressed temperature effects on CH₄ oxidation and methanotrophic communities, there is little knowledge about the physiological adjustments that underlie these effects. We have studied thermal acclimation in Methylobacter, a widespread, abundant, and environmentally important methanotrophic genus. Comparisons of growth and CH₄ oxidation kinetics at different temperatures in three members of the genus demonstrate that temperature has a strong influence on how much CH₄ is consumed to support growth at different CH₄ concentrations. However, the temperature effect varies considerably between species, suggesting that how a methanotrophic community is composed influences the temperature effect on CH₄ uptake. To understand thermal acclimation mechanisms widely we carried out a transcriptomics experiment with Methylobacter tundripaludum SV96^T. We observed, at different temperatures, how varying abundances of transcripts for glycogen and protein biosynthesis relate to cellular glycogen and ribosome concentrations. Our data also demonstrated transcriptional adjustment of CH₄ oxidation, oxidative phosphorylation, membrane fatty acid saturation, cell wall composition, and exopolysaccharides between temperatures. In addition, we observed differences in M. tundripaludum SV96^T cell sizes at different temperatures. We conclude that thermal acclimation in Methylobacter results from transcriptional adjustment of central metabolism, protein biosynthesis, cell walls and storage. Acclimation leads to large shifts in CH₄ consumption and growth efficiency, but with major differences between species. Thus, our study demonstrates that physiological adjustments to temperature change can substantially influence environmental CH₄ uptake rates and that consideration of methanotroph physiology might be vital for accurate predictions of warming effects on CH₄ emissions.