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Modulating Pathway Performance by Perturbing Local Genetic Context
journal contribution
posted on 2020-03-24, 15:36 authored by Carmen Lopez, Yuxin Zhao, Rick Masonbrink, Zengyi ShaoCombinatorial
engineering is a preferred strategy for attaining
optimal pathway performance. Previous endeavors have been concentrated
on regulatory elements (e.g., promoters, terminators,
and ribosomal binding sites) and/or open reading frames. Accumulating
evidence indicates that noncoding DNA sequences flanking a transcriptional
unit on the genome strongly impact gene expression. Here, we sought
to mimic the effect imposed on expression cassettes by the genome.
We created variants of the model yeast Saccharomyces cerevisiae with significantly improved fluorescence or cellobiose consumption
rate by randomizing the sequences adjacent to the GFP expression cassette
or the cellobiose-utilization pathway, respectively. Interestingly,
nucleotide specificity was observed at certain positions and showed
to be essential for achieving optimal cellobiose assimilation. Further
characterization suggested that the modulation effects of the short
sequences flanking the expression cassettes could be potentially mediated
by remodeling DNA packaging and/or recruiting transcription factors.
Collectively, these results indicate that the often-overlooked contiguous
DNA sequences can be exploited to rapidly achieve balanced pathway
expression, and the corresponding approach could be easily stacked
with other combinatorial engineering strategies.