Epigenetics as a design variable in heterologous expression

Heterologous expression-the expression of genes from one organism into another-is an important approach to produce value-added chemicals such as proteins and metabolites. The main challenge, however, is achieving high production yields that are commercially viable. Approaches used to improve productivity rely on editing the DNA sequence, including promoters, ribosome binding sites, and codon usage. Epigenetics, in the form of DNA methylation, may offer an additional layer of control to improve production yields but remains unexplored. Moreover, the choice of the host organism is crucial. We have been interested in developing the alternative bacterial host Burkholderia sp. FERM BP-3421. Four DNA methyltransferases were predicted to be encoded in the genome of this strain, one of which was silent under our growth conditions. After performing gene deletions or overexpression, we experimentally assigned the recognition motif of each DNA methyltransferase, one of which was shown to be essential. As expected, modulation of DNA methylation had major impacts on the transcriptome and metabolome of the host, varying depending on which DNA methyltransferase was modified. Two of the methyltransferases that methylate adenine had overlapping profiles, whereas the one methylating cytosine differed. Finally, we probed the role of DNA methylation during heterologous expression using green fluorescence protein and three different promoters. Expression levels varied in a context-dependent manner with mechanisms, in some cases, involving the presence of methylation motifs within promoter regions but also based on global host effects. This work establishes epigenetics as an important design variable to help improve heterologous expression yields.IMPORTANCEImproving heterologous expression currently relies on modifying the DNA sequence. Epigenetics offers an opportunity to improve outcomes but remains unexplored. We first investigated the effect of epigenetic DNA methylation on the growth, transcriptome, and metabolome of an alternative bacterial host we are interested in developing by generating three single, DNA methyltransferase mutants. Next, we showed that epigenetic DNA methylation or the lack thereof can affect heterologous expression outcomes. The effect was context-dependent, reaching 2.4-fold improvement in one instance. This work highlights the importance of being epigenetics-aware during host development for heterologous expression.