In:
Science, American Association for the Advancement of Science (AAAS), Vol. 378, No. 6624 ( 2022-12-09)
Kurzfassung:
The intracellular concentration of a protein depends on the rates of several processes, including transcription, translation, and the degradation and/or dilution of messenger RNAs (mRNAs) and proteins. These rates can be vastly different for different genes and across different growth conditions because of gene-specific regulation. At the systems level, protein concentrations are further affected by the availability of shared gene expression machineries—e.g., RNA polymerases and ribosomes—and are constrained by the approximately invariant cellular mass density. Even in one of the best-characterized model organisms, Escherichia coli , it is unclear how the gene-specific and systems-level effects work together toward setting the cellular proteome. This knowledge gap has not only hindered our efforts in building a predictive framework of gene expression but has also limited our abilities in guiding the rational design of gene circuits. RATIONALE We undertook a quantitative, genome-scale study, combining experimental and theoretical approaches, to tease apart the contribution of the specific and global effects on cellular protein concentrations in exponentially growing E. coli cells across a variety of growth conditions. We complemented genome-scale proteomic and transcriptomic data with biochemical measurements of total absolute mRNA abundances and synthesis rates. We compared these measurements to gene dosage and the concentrations of ribosomes and RNA polymerases to quantitatively characterize the activity of the gene expression machinery across conditions. This comprehensive dataset allowed us to analyze, in quantitative detail, the interplay between the activity of gene expression machinery, the activity of individual promoters, and the resulting protein concentrations. RESULTS We compiled a comprehensive atlas of the determinants of gene expression across conditions—from the concentrations of genes, mRNAs, and proteins to the rates of transcriptional and translational initiation and mRNA degradation for thousands of genes. We were able to determine the on rate of each promoter, a quantity capturing the overall effect of transcriptional regulation that has been elusive through most existing gene expression studies. Unexpectedly, we found that for most genes, the cytosolic protein concentrations were primarily determined by the innate magnitude of their promoter on rates, which spanned more than three orders of magnitude. Changes in protein concentrations resulting from changes in growth conditions were typically much smaller—well within one order of magnitude—and were mostly exerted through changes in transcription initiation. E. coli ’s strategy to implement gene regulation can be summarized by two design principles. First, protein concentrations are predominantly set transcriptionally, with relatively invariant posttranscriptional characteristics (translation efficiencies and degradation rates) for most mRNAs and growth conditions. Second, the overall fluxes of transcription and translation are tightly coordinated: The average density of five ribosomes per kilobase is nearly invariant across mRNA species and across growth conditions, even though the mRNA and ribosome abundances can each vary substantially. We find this coordination to be implemented through the anti-sigma factor Rsd, which modulates the availability of RNA polymerases for transcription across different growth conditions. These two principles lead to a quantitative formulation of the central dogma of bacterial gene expression, connecting mRNA and protein concentrations to the regulatory activities of the corresponding promoters. CONCLUSION These quantitative relationships reveal the unexpectedly simple strategies used by E. coli to attain desired protein concentrations despite the complexity of global physiological constraints: Individual protein concentrations are primarily set by gene-specific transcriptional regulation, with global transcriptional regulation set to cancel the strong growth rate dependence of protein synthesis. These relations provide the basis for understanding the behavior of more complex genetic circuits in different conditions and for the inverse problem of deducing regulatory activities given the observed mRNA and protein levels. Principles governing gene expression in E. coli . The availability of RNA polymerases (RNAPs) is coordinated with that of ribosomes, and the mRNA characteristics (translation efficiency and degradation rates) are uniform across most genes and growth conditions. These two principles prescribe a simple gene expression strategy in which protein concentrations are almost entirely controlled at the promoter level for most genes.
Materialart:
Online-Ressource
ISSN:
0036-8075
,
1095-9203
DOI:
10.1126/science.abk2066
Sprache:
Englisch
Verlag:
American Association for the Advancement of Science (AAAS)
Publikationsdatum:
2022
ZDB Id:
128410-1
ZDB Id:
2066996-3
ZDB Id:
2060783-0
SSG:
11
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