Synthetic Biology Parts, Devices and Applications

110 6 Constitutive and Regulated Promoters in Yeast

In this chapter, we deal with natural and synthetic promoters frequently used

in yeast. After giving an overview of the essential features of natural promoters,

we describe principles and strategies exploited to produce synthetic promoters

and their cognate transcription factors. We leave out from the discussion other

aspects of gene expression regulation, like gene copy number, transcription elon-

gation and termination, transcript processing, mRNA stability, translation, and

protein stability.

6.2 Yeast Promoters

A promoter is a DNA sequence enabling and regulating transcription initiation.

In this section, we point out the essential structural and functional features

of  yeast promoters. For more detailed descriptions, reviews are available

[10–14].

Yeast promoters consist of two functionally and physically distinguishable

regions: the core promoter and the upstream element [4, 15, 16] (Figure 6.1,

top). The core promoter is the region that carries the minimal information

needed to start transcription, independently of any regulation [3, 15, 17]. It

NN NN NN NNNNN NNN

Point mutate Substitute non-consensus with

random oligonucleotides

TFBSs TATA TSS

Upstream elementCore promoter

ORFORF

Figure 6.1 Modified natural yeast promoters. Top, typical bipartite structure of yeast

promoters. Bottom left, promoter libraries obtained by point mutation. Random point

mutations, illustrated as stars, are introduced by error-prone PCR along the sequence of the

starting promoter. Bottom right, promoter libraries obtained by substituting non-consensus

sequences with random oligonucleotides. By concentrating the mutations in the non-

consensus regions, it is possible to fine-tune the strength of the starting promoter. N:

nucleotide; ORF: open reading frame; TATA: TATA element; TFBS: transcription factor binding

site; TSS: transcription initiation start site.