Just-in-time transcription program in metabolic pathways

Abstract

A primary goal of systems biology is to understand the design principles of the transcription networks that govern the timing of gene expression 1-5 . Here we measured promoter activity for ∼100 genes in parallel from living cells at a resolution of minutes and accuracy of 10%, based on GFP and Lux reporter libraries 3 . Focusing on the amino-acid biosynthesis systems of Escherichia coli 4 , we identified a previously unknown temporal expression program and expression hierarchy that matches the enzyme order in unbranched pathways. We identified two design principles: the closer the enzyme is to the beginning of the pathway, the shorter the response time of the activation of its promoter and the higher its maximal promoter activity. Mathematical analysis suggests that this 'just-in-time' (ref. 5) transcription program is optimal under constraints of rapidly reaching a production goal with minimal total enzyme production 6,7 . Our findings suggest that metabolic regulation networks are designed to generate precision promoter timing and activity programs that can be understood using the engineering principles of production pipelines.Amino-acid biosynthesis (AAB) in E. coli is carried out by well-characterized enzymatic pathways 4,6-11 . The genes encoding these enzymes are governed by a transcriptional regulatory network 12,13 , which is an excellent model system for studying the design principles of metabolic regulation. To study the dynamics of transcription of AAB genes at high temporal resolution and accuracy, we constructed a library of 52 reporter strains that represent ∼50% of known AAB genes. We designed each reporter strain by cloning one of the promoter regions of E. coli K-12 MG1655 upstream of a Lux or a fast-folding GFP reporter gene (Fig. 1a). We measured promoter activity with a high temporal resolution by measuring fluorescence, luminescence and absorbance from 96 cultures in parallel in a multiwell fluorimeter 3,14 .

Activation of the Escherichia coli marA/soxS/rob Regulon in Response to Transcriptional Activator Concentration

“…13,21 Furthermore, because promoters of the regulon control expression of functionally diverse genes, the temporal ordering might not lead to the advantages in efficiency proposed for temporal ordering of functionally coherent regulons. [18][19][20] We also note that the first promoter activated by either activator is marRAB itself. 21 This has two consequences: 1) MarA autocatalytically increases its own synthesis, which is a rare feature among transcription factors in E. coli.…”

How Salmonella typhimurium measures the length of flagellar filaments

“…A flagellumspecific sigma factor in the second class regulates the expression of other operons in the same or lower classes. The basal structure and hook must be complete before the proteins for the filament are expressed, so that transcription of the filament components is "just-in-time" [11].…”

Evolution of transcription factor DNA binding sites

“…Some genes should be under highly sensitive regulation, others by weaker regulation, and the efficiency of regulation is determined by binding affinity of the regulatory factor (Berg and von Hippel, 1987). For instance, recently it has been shown, that the closer the enzyme is to the beginning of the pathway, the shorter is the response time of the activation of its promoter and the higher is its maximal promoter activity (Zaslaver et al, 2004). In this model, the consensus nucleotide provides the maximal binding site affinity, which is required, however, only for a subset of all genes.…”