Cell-free gene-regulatory network engineering with synthetic transcription factors.

Abstract:

:Gene-regulatory networks are ubiquitous in nature and critical for bottom-up engineering of synthetic networks. Transcriptional repression is a fundamental function that can be tuned at the level of DNA, protein, and cooperative protein-protein interactions, necessitating high-throughput experimental approaches for in-depth characterization. Here, we used a cell-free system in combination with a high-throughput microfluidic device to comprehensively study the different tuning mechanisms of a synthetic zinc-finger repressor library, whose affinity and cooperativity can be rationally engineered. The device is integrated into a comprehensive workflow that includes determination of transcription-factor binding-energy landscapes and mechanistic modeling, enabling us to generate a library of well-characterized synthetic transcription factors and corresponding promoters, which we then used to build gene-regulatory networks de novo. The well-characterized synthetic parts and insights gained should be useful for rationally engineering gene-regulatory networks and for studying the biophysics of transcriptional regulation.

authors

Swank Z,Laohakunakorn N,Maerkl SJ

doi

10.1073/pnas.1816591116

subject

Has Abstract

pub_date

2019-03-26 00:00:00

pages

5892-5901

issue

13

eissn

0027-8424

issn

1091-6490

pii

1816591116

journal_volume

116

pub_type

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