Methanol regulated yeast promoters: production vehicles and toolbox for synthetic biology
© Gasser et al. 2015
Received: 19 November 2015
Accepted: 25 November 2015
Published: 2 December 2015
Promoters are indispensable elements of a standardized parts collection for synthetic biology. Regulated promoters of a wide variety of well-defined induction ratios and expression strengths are highly interesting for many applications. Exemplarily, we discuss the application of published genome scale transcriptomics data for the primary selection of methanol inducible promoters of the yeast Pichia pastoris (Komagataella sp.). Such a promoter collection can serve as an excellent toolbox for cell and metabolic engineering, and for gene expression to produce heterologous proteins.
A major task of synthetic biology is the provision of standardized elements for rapid assembly of predictable recombinant gene expression cassettes [1, 2]. These elements include vectors, selection markers, and most importantly collections of regulatory elements like promoters, transcription terminators, secretory leaders and other signal sequences. Ideally, collections of these parts are cataloged in standardized, easy to assemble formats like BioBrick . Promoters are indispensable parts for synthetic biology approaches  and are needed for different expression strength in order to balance the expression levels in a synthetic pathway . There are a plethora of studies which characterize, e.g. constitutive promoters of different strength for Escherichia coli , Aspergillus niger  or Pichia pastoris . Depending on the application it might be necessary to tightly control the promoter activity. Especially regulated promoters are often strictly host specific, so that they need to be identified, characterized and standardized for the host species of interest, as shown e.g. for E. coli .
Methanol regulated promoters
Methylotrophic yeasts such as P. pastoris (syn. Komagataella sp.) have gained great interest as production hosts for recombinant proteins  and more recently also as platform for metabolite production . Both applications require promoter collections of different strength for metabolic and cell engineering to enable and enhance productivity. Promoter libraries were developed based on mutating transcription factor binding sites , or by random mutagenesis . Strong constitutive and regulated promoters were identified by transcriptomics studies [12, 13]. Delic et al.  described a collection of native regulated promoters of different strength with the main aim of providing repressible promoters for gene knockdown studies. Synthetic core promoters represent a source for transcriptional initiators at different strength, however with the loss of regulatory features [1, 15].
Methanol regulated genes of P. pastoris as a source of regulated promoters
Ranked expression level (methanol)a
Co-regulation: 1 = with A/D/F; 2 = with A; 3 = with D/F; 4 = up at glucose limitc
Genome scale transcriptomic studies are a valuable source of information on native promoters and have been successfully used to identify promoters of different strength and desired regulatory behavior. Well defined promoters are core elements of synthetic biology part collections. The collection of P. pastoris promoters presented here, and others analyzed in the cited references can serve as a basis for setting up a P. pastoris promoter collection. Promoters with different regulatory strength are crucial elements of toolboxes for cell and metabolic engineering. In addition, they can be directly employed for gene expression to produce heterologous proteins or metabolites in yeasts.
All authors contributed equally to this commentary. All authors read and approved the final manuscript.
The authors declare that they have no competing interests.
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