FRINK Linked Data Fragments server

Matches in SemOpenAlex for { <https://semopenalex.org/work/W2175978653> ?p ?o ?g. }

• W2175978653 abstract "Article23 November 2015Open Access Source Data A synthetic growth switch based on controlled expression of RNA polymerase Jérôme Izard Jérôme Izard Université Grenoble Alpes, Laboratoire Interdisciplinaire de Physique (CNRS UMR 5588), Saint Martin d'Hères, France INRIA, Grenoble – Rhône-Alpes research center, Saint Ismier, France These authors contributed equally to this work Search for more papers by this author Cindy DC Gomez Balderas Cindy DC Gomez Balderas Université Grenoble Alpes, Laboratoire Interdisciplinaire de Physique (CNRS UMR 5588), Saint Martin d'Hères, France INRIA, Grenoble – Rhône-Alpes research center, Saint Ismier, France These authors contributed equally to this work Search for more papers by this author Delphine Ropers Delphine Ropers INRIA, Grenoble – Rhône-Alpes research center, Saint Ismier, France Search for more papers by this author Stephan Lacour Stephan Lacour Université Grenoble Alpes, Laboratoire Interdisciplinaire de Physique (CNRS UMR 5588), Saint Martin d'Hères, France INRIA, Grenoble – Rhône-Alpes research center, Saint Ismier, France Search for more papers by this author Xiaohu Song Xiaohu Song Center for Research and Interdisciplinarity, INSERM U1001, Medicine Faculty, Site Cochin Port-Royal, University Paris Descartes, Paris, France Search for more papers by this author Yifan Yang Yifan Yang Center for Research and Interdisciplinarity, INSERM U1001, Medicine Faculty, Site Cochin Port-Royal, University Paris Descartes, Paris, France Search for more papers by this author Ariel B Lindner Ariel B Lindner Center for Research and Interdisciplinarity, INSERM U1001, Medicine Faculty, Site Cochin Port-Royal, University Paris Descartes, Paris, France Search for more papers by this author Johannes Geiselmann Corresponding Author Johannes Geiselmann Université Grenoble Alpes, Laboratoire Interdisciplinaire de Physique (CNRS UMR 5588), Saint Martin d'Hères, France INRIA, Grenoble – Rhône-Alpes research center, Saint Ismier, France These authors contributed equally to this work Search for more papers by this author Hidde de Jong Corresponding Author Hidde de Jong INRIA, Grenoble – Rhône-Alpes research center, Saint Ismier, France These authors contributed equally to this work Search for more papers by this author Jérôme Izard Jérôme Izard Université Grenoble Alpes, Laboratoire Interdisciplinaire de Physique (CNRS UMR 5588), Saint Martin d'Hères, France INRIA, Grenoble – Rhône-Alpes research center, Saint Ismier, France These authors contributed equally to this work Search for more papers by this author Cindy DC Gomez Balderas Cindy DC Gomez Balderas Université Grenoble Alpes, Laboratoire Interdisciplinaire de Physique (CNRS UMR 5588), Saint Martin d'Hères, France INRIA, Grenoble – Rhône-Alpes research center, Saint Ismier, France These authors contributed equally to this work Search for more papers by this author Delphine Ropers Delphine Ropers INRIA, Grenoble – Rhône-Alpes research center, Saint Ismier, France Search for more papers by this author Stephan Lacour Stephan Lacour Université Grenoble Alpes, Laboratoire Interdisciplinaire de Physique (CNRS UMR 5588), Saint Martin d'Hères, France INRIA, Grenoble – Rhône-Alpes research center, Saint Ismier, France Search for more papers by this author Xiaohu Song Xiaohu Song Center for Research and Interdisciplinarity, INSERM U1001, Medicine Faculty, Site Cochin Port-Royal, University Paris Descartes, Paris, France Search for more papers by this author Yifan Yang Yifan Yang Center for Research and Interdisciplinarity, INSERM U1001, Medicine Faculty, Site Cochin Port-Royal, University Paris Descartes, Paris, France Search for more papers by this author Ariel B Lindner Ariel B Lindner Center for Research and Interdisciplinarity, INSERM U1001, Medicine Faculty, Site Cochin Port-Royal, University Paris Descartes, Paris, France Search for more papers by this author Johannes Geiselmann Corresponding Author Johannes Geiselmann Université Grenoble Alpes, Laboratoire Interdisciplinaire de Physique (CNRS UMR 5588), Saint Martin d'Hères, France INRIA, Grenoble – Rhône-Alpes research center, Saint Ismier, France These authors contributed equally to this work Search for more papers by this author Hidde de Jong Corresponding Author Hidde de Jong INRIA, Grenoble – Rhône-Alpes research center, Saint Ismier, France These authors contributed equally to this work Search for more papers by this author Author Information Jérôme Izard1,2, Cindy DC Gomez Balderas1,2, Delphine Ropers2, Stephan Lacour1,2, Xiaohu Song3, Yifan Yang3, Ariel B Lindner3, Johannes Geiselmann 1,2 and Hidde Jong 2 1Université Grenoble Alpes, Laboratoire Interdisciplinaire de Physique (CNRS UMR 5588), Saint Martin d'Hères, France 2INRIA, Grenoble – Rhône-Alpes research center, Saint Ismier, France 3Center for Research and Interdisciplinarity, INSERM U1001, Medicine Faculty, Site Cochin Port-Royal, University Paris Descartes, Paris, France *Corresponding author. Tel: +33 476514753; E-mail: [email protected] *Corresponding author. Tel: +33 476615335; E-mail: [email protected] Molecular Systems Biology (2015)11:840https://doi.org/10.15252/msb.20156382 PDFDownload PDF of article text and main figures. Peer ReviewDownload a summary of the editorial decision process including editorial decision letters, reviewer comments and author responses to feedback. ToolsAdd to favoritesDownload CitationsTrack CitationsPermissions Figures & Info Abstract The ability to control growth is essential for fundamental studies of bacterial physiology and biotechnological applications. We have engineered an Escherichia coli strain in which the transcription of a key component of the gene expression machinery, RNA polymerase, is under the control of an inducible promoter. By changing the inducer concentration in the medium, we can adjust the RNA polymerase concentration and thereby switch bacterial growth between zero and the maximal growth rate supported by the medium. We show that our synthetic growth switch functions in a medium-independent and reversible way, and we provide evidence that the switching phenotype arises from the ultrasensitive response of the growth rate to the concentration of RNA polymerase. We present an application of the growth switch in which both the wild-type E. coli strain and our modified strain are endowed with the capacity to produce glycerol when growing on glucose. Cells in which growth has been switched off continue to be metabolically active and harness the energy gain to produce glycerol at a twofold higher yield than in cells with natural control of RNA polymerase expression. Remarkably, without any further optimization, the improved yield is close to the theoretical maximum computed from a flux balance model of E. coli metabolism. The proposed synthetic growth switch is a promising tool for gaining a better understanding of bacterial physiology and for applications in synthetic biology and biotechnology. Synopsis Bacterial growth can be switched on/off by expressing the RNA polymerase large subunits under the control of an inducible promoter. When growth is switched off, cells reorient nutrient fluxes to increase the production of a metabolite of interest. An E. coli strain is constructed in which the rpoBC genes, encoding the ββ′ subunits of RNA polymerase, are placed under the control of an IPTG-inducible promoter. The growth of the strain can be switched on and off by varying the concentration of IPTG in the medium. The growth switch is reversible and does not depend on the specific nutrient composition of the medium. A proof-of-principle of the biotechnological relevance of the growth switch is provided by showing that growth-arrested cells are capable of converting glucose to glycerol at a yield that is close to the theoretical maximum. Introduction Optimizing growth in a given environment is a major challenge for microorganisms. In particular, changes in nutrient availability require a rapid adjustment of growth rate and therefore the reallocation of available resources to cellular functions. Early work in bacterial physiology has characterized the macromolecular composition of microbial cells as a function of growth rate (Schaechter et al, 1958; Kjeldgaard 1961; Neidhardt & Fraenkel, 1961; Bremer & Dennis, 1996; reviewed in Maaløe & Kjeldgaard, 1966; Scott & Hwa, 2011). These experiments have shown that the DNA, RNA, and protein contents are correlated with the growth rate and not the nature of the specific nutrients available for growth. The culture history also plays a role, especially when the nutritional quality of the medium is low (Ehrenberg et al, 2013). Other physiological quantities that vary with the growth rate include the intracellular concentrations of RNA polymerase and ribosome, the mass ratio of total protein and RNA, and the size of metabolic pools (Bremer & Dennis, 1996). Recently, there has been a regained interest in the role of global cell physiology, notably the activity of the transcriptional and translational machinery, in the control of gene expression, both in balanced growth and during growth transitions (Klumpp et al, 2009; Levy & Barkai, 2009; Molenaar et al, 2009; Scott et al, 2010; Goelzer & Fromion, 2011; Karr et al, 2012; Berthoumieux et al, 2013; Ehrenberg et al, 2013; Gerosa et al, 2013; Keren et al, 2013; O'Brien et al, 2013; You et al, 2013; de Lorenzo, 2014; Scott et al, 2014; Weisse et al, 2015). The ability to experimentally control the growth rate is thus crucial for studying bacterial physiology. It is also of central importance for applications in biotechnology, where often the goal is to limit or even arrest growth. Growth-arrested cells with a functional metabolism open the possibility to channel resources into the production of a desired metabolite, instead of wasting nutrients on biomass production (Flickinger & Rouse, 1993; Sonderegger et al, 2005). A variety of approaches have been classically employed to limit growth. One example is the use of antibiotics targeting the transcription or translation machinery or DNA topology. This approach can lead to cell death, however, and in addition to being too costly in an industrial setting, mutants resistant to antibiotics quickly emerge and take over the population. An alternative strategy consists in imposing limitations on nutrients essential for growth, in particular carbon, nitrogen, and phosphate sources. This intervention strongly affects metabolism though, as cells adjust their flux distribution and enzyme levels to nutrient limitations. As a consequence, the engineered metabolic pathways and genetic circuits may not function or function suboptimally. The above limitations call for novel strategies to limit or completely stop growth. Ideally, it should be possible to grow a cell population to a certain biomass and then switch off growth. The enzymes present at the time of growth arrest are expected to remain functional, thus enabling high-yield production of a metabolite of interest. When the degradation of enzymes and other proteins threatens the stability of metabolism, it should be possible to switch on growth again, thus alternating phases of biomass accumulation and product synthesis. In addition to being reversible, the exercised control over the growth rate should be medium independent, in the sense of being applicable over a wide range of medium compositions and corresponding nutrient uptake patterns. Here, we present a synthetic growth switch that satisfies the above criteria. We report the construction of an Escherichia coli strain in which the transcription of the operon encoding two large subunits of RNA polymerase, β and β′, is under the tight control of an isopropyl-β-D-1-thiogalactopyranoside (IPTG)-inducible promoter. Previous attempts at constructing such a strain have failed to give conclusive results (Nomura et al, 1987). The challenge has been abandoned after the heyday of research in bacterial transcription, but modern techniques of chromosome engineering have allowed us to successfully tackle this endeavor. In our engineered strain, the growth rate can be switched on and off by changing the inducer concentration in the medium and thus the concentration of RNA polymerase. The system is purely genetic and therefore does not depend on the composition of the medium, apart from the supply of inducer. When the production of RNA polymerase is completely shut off, the growth rate decreases continuously, but cells remain metabolically active and do not die. The analysis of single cells in a microfluidics device shows that growth control is reversible: a growth-arrested culture resumes normal growth after the inducer, IPTG, is added back to the medium. Our data suggest that the switch between growth and growth arrest is due to the highly ultrasensitive response of the growth rate to a change in concentration of the β and β′ subunits, as quantified by means of a fluorescent tag on β′. This indicates that the variation of rpoBC expression is the effective cause of the change in growth rate. In order to provide a proof-of-principle of the applicability of our growth switch in a biotechnological context, we endow our E. coli strain with the capacity to convert glucose to glycerol, by expressing appropriate yeast enzymes. We measure extracellular levels of glucose and glycerol and show that this allows the production of glycerol during growth arrest, at a yield much higher than that obtained in a growing wild-type strain expressing the same enzymes. Remarkably, the glycerol production yield is close to the maximum theoretical yield computed from a flux balance model of E. coli metabolism. These results demonstrate that the desired reallocation of resources from growth toward the production of a molecule of interest is feasible and indeed highly profitable. In summary, we present a growth switch that is a generic tool of broad interest for applications in synthetic biology and biotechnology. Since the controller is a genetic element, our system is modular: the growth switch can be interfaced with any other (natural or synthetic) cellular network via intracellular signals. Moreover, the principle underlying its design is applicable to other bacteria with a single RNA polymerase. Results Reengineering the control of RNA polymerase expression The core RNA polymerase of Escherichia coli consists of three different subunits and has the composition ′. The α subunit being produced in excess during growth (Hayward & Fyfe, 1978), the amount of the ββ′ subunits sets the overall concentration of RNA polymerase (Engbaek et al, 1976; Kawakami et al, 1979; Shepherd et al, 2001). These two subunits are the product of the rpoB and rpoC genes, organized in an operon with upstream genes coding for ribosomal proteins (rplKAJL) (Yamamoto & Nomura, 1978). The expression of rpoBC genes is regulated both at the transcriptional and translational level by complex and still incompletely understood mechanisms. An attenuator of transcription located in the intergenic region rplL-rpoB decouples the expression of ribosomal proteins and the two RNA polymerase subunits (Ishihama & Fukuda, 1980; Dykxhoorn et al, 1996). Growth of a bacterial cell requires the duplication of its contents, in particular protein, RNA, and DNA which make up 75–90% of cellular material (Bremer & Dennis, 1996). Synthesis of RNA and protein starts with transcription, so the production of RNA polymerase is essential for growth and survival of the cell. When the activity of RNA polymerase is inhibited, for example, by the action of an antibiotic such as rifampicin, the bacteria stop growing (Campbell et al, 2001). Since E. coli possesses only one RNA polymerase, this suggests that, by adjusting the concentration of the limiting β and β′ subunits, we can vary the growth rate of the cell. We therefore replaced the natural promoter of rpoBC of the E. coli strain BW25113 by an IPTG-inducible promoter. The transcription of rpoBC was completely isolated from the upstream ribosomal proteins by introducing a selection cassette and a strong transcriptional terminator (Fig 1A and Appendix Fig S1). Two extra copies of the Lac repressor gene were added to the chromosome in two different loci in order to prevent the relief of repression by mutations inactivating the single original copy of lacI (Fig 1B and Appendix Fig S2). Whereas a bacterial population (typically between and bacteria in a growth experiment) most likely contains an individual carrying a mutation in one copy of lacI, a double mutation that inactivates both copies of the gene is very unlikely. Hereafter, we call the resulting strain with reengineered transcriptional control of RNA polymerase expression the “R strain” and the wild-type strain from which it has been derived the “W strain”. Figure 1. Construction of an E. coli strain with inducible expression of the rpoBC genes encoding the ββ′ subunits of RNA polymerase We have replaced the rpoBC promoter region by an IPTG-inducible promoter. A strong transcriptional terminator, rrnBt1, was inserted upstream of the selection cassette (spcR) and the T5 promoter with two lac operator sequences. rpoBC transcription is thus efficiently repressed by the lac repressor LacI (Lutz & Bujard, 1997). The gene encoding LacI is present in three copies on the chromosome, one natural copy (in black) and two additional copies (in red). The engineered strain is referred to as “R”, in distinction to the wild-type strain “W”. For more details on the construction, see Appendix Figs S1 and S2. Download figure Download PowerPoint Reengineering transcriptional control of RNA polymerase allows a medium-independent growth switch Does the strain with inducible transcriptional control of the RNA polymerase β and β′ subunits indeed allow the growth rate to be controlled? In order to answer this question, we characterized the R strain in different growth media, adjusting rpoBC expression by varying the IPTG concentration. The R and W strains were incubated on LB agar plates with a high concentration of IPTG (1,000 μM) and a single colony was picked from the plates and precultured in M9 minimal medium, supplemented with 0.2% glucose and 1,000 μM IPTG to ensure maximal production of RNA polymerase. The overnight precultures were centrifuged, washed, and rediluted into 96-well microplates containing fresh M9 minimal medium with 0.2% glucose and different concentrations of IPTG (4). The microplate culture conditions were chosen so as to improve aeration (use of glass beads for stirring, high frequency of shaking) and obtain the same growth rate as in shake flasks (see below). Figure 2A shows the growth curves of the R strain obtained for a range of IPTG concentrations, varying from 0 to 1,000 μM. As a control, we also show the growth curve of the W strain. The growth curves are identical for all concentrations of IPTG during the first 3 h of the experiment because there is enough RNA polymerase from the overnight preculture to ensure a maximal growth rate. After this initial growth phase determined by the conditions of the preculture, the concentration of IPTG controls growth of the R strain in a dose-dependent manner. To a first approximation, the growth curves can be divided into two categories. In the first category, for IPTG concentrations of 30 μM and higher, growth is normal or close to normal, as compared to the wild-type strain, and the cultures reach stationary phase at an absorbance of 0.5, once all nutrients in the medium have been exhausted. The bacteria first consume all glucose in the medium and then continue growth at a lower rate, utilizing by-products secreted during growth on glucose, notably acetate (Andersen & von Meyenburg, 1980; El-Mansi & Holms, 1989; Wolfe, 2005). For the highest concentrations of IPTG (100 and 1,000 μM), the growth kinetics of the R strain is practically indistinguishable from that of the W strain. In the second category, for IPTG concentrations of 20 μM and lower, growth stops after a few hours at an absorbance level well inferior to that reached in the wild-type strain. Figure 2. External control of the expression of rpoBC yields a growth switch in E. coli The growth kinetics of the R strain changes as a function of the concentration of IPTG added to M9 minimal medium supplemented with 0.2% glucose. Blue curves are averaged absorbance measurements at 600 nm (five biological replicates), for increasing concentrations of IPTG (0, 10, 20, 30, 40, 50, 100, 1,000 μM). Shaded areas represent ± two standard errors of the mean. The red curve represents the growth kinetics of the W strain. The growth rates are typically computed in the time intervals indicated by the green bars. The growth rate of the R strain responds in a switch-like manner to IPTG addition, with a threshold between 20 and 30 μM IPTG. The figure shows the growth rate estimated from the absorbance data, as explained in the main text, for both the R strain (blue) and the W strain (red). The reported growth rate is the mean over five replicates and the error bars are defined as ± two standard errors of the mean. The black circles represent the growth rates obtained with the R and W strains in shake flask experiments, which are in very good agreement with those obtained in microplate experiments. The dashed curve is obtained by fitting a Michaelis–Menten function to the data above the IPTG threshold and setting its value to 0 below the threshold. Idem for M9 medium supplemented with 0.2% glucose in R-ΔlacY strain. Idem for M9 minimal medium supplemented with 0.2% glucose and 0.2% casamino acids. Idem for LB medium supplemented with 0.2% glucose. Comparison of the growth-rate response of the R strain to different concentrations of IPTG. In all conditions, a switch occurs, in the sense that the growth rate reacts abruptly to a small increase in IPTG just above the threshold. The R-ΔlacY strain behaves quite similarly to the R strain in the reference medium (M9 medium with 0.2% glucose), indicating that the growth switch is not due to the cooperativity of the lac induction system. Source data are available online for this figure. Source Data for Figure 2 [msb156382-sup-0005-SDataFig2.zip] Download figure Download PowerPoint In order to quantify these observations, we computed the growth rate attained for the different concentrations of IPTG at an appropriate stage of the growth kinetics. More precisely, for concentrations of 20 μM and lower, we computed the growth rate after growth arrest at low absorbance (beyond 1,000 min). For concentrations of 30 μM and higher, we considered the time window where the absorbance is larger than 0.05 (after 5–6 generations, when RNA polymerase from the preculture has been strongly diluted out) and smaller than 0.2 (above which growth is no longer exponential, due to growth-limiting oxygen transfer rates). In every case, we fitted an exponential function to the data, typically consisting of several dozens of absorbance measurements, to obtain a precise estimate of the growth rate. The results in Fig 2B provide a quantitative picture of the observed growth switch. At IPTG concentrations of 20 μM and lower, the growth rate is 0, while between 20 and 30 μM it jumps to values beyond 0.008 min. For the highest IPTG concentrations, the growth rate takes a value of 0.012 ± 0.0008 min, corresponding to a doubling time of 57 min, characteristic for the wild-type BW25113 E. coli strain (Volkmer & Heinemann, 2011). For IPTG concentrations of 30 μM and higher, we repeated the experiment with a 10-fold higher dilution ratio of the preculture, so as to make sure that the observed depletion of the nutrients in the medium, and the corresponding growth rate, is really due to the continued synthesis of RNA polymerase at the specified IPTG concentration and not a residual effect from the preculture (Fig EV1). We also compared the growth rate computed from the data obtained from microplate experiments with that obtained in experiments in shake flasks and observed excellent agreement (Fig 2B). This assures that the growth conditions in the microplate, at least for the bacterial population densities considered here, are sufficiently similar to growth conditions in shake flasks. Furthermore, we tested that the growth rate of the W strain did not vary with the IPTG concentrations, showing that the effect of IPTG on the growth rate is really due to transcription regulation of the rpoBC genes and not to some other side-effect (Fig EV1). Click here to expand this figure. Figure EV1. Growth rate and mCherry concentration in additional strains and conditionsAdditional controls of the IPTG dependence of the growth rate in the W and R strains, with and without the mCherry tag on rpoC. All experiments have been carried out in the conditions summarized in Fig 2 including the number of replicates and data treatment. The growth rate of the W strain in M9 medium with 0.2% glucose does not vary with the concentration of IPTG. Variation of the growth rate with IPTG concentration in the R strain grown in M9 minimal medium with 0.2% succinate, using the W strain as a control. The maximum growth rate attained is 0.009 ± 0.0007 min, corresponding to a doubling time of 77 min. 10 μM IPTG is sufficient for growth, consistent with the lower growth rate supported by this medium in comparison with the reference medium, M9 minimal medium supplemented with glucose. Variation of the growth rate with the IPTG concentration in the R-rpoC-mCherry strain grown in M9 medium with 0.2% glucose, with the W-rpoC-mCherry strain as a control (in red). The switching phenotype is the same as in the R strain. The concentration of mCherry as a function of IPTG in the W-rpoC-mCherry strain, grown in M9 medium with 0.2% glucose. In a strain in which rpoBC expression is controlled by the natural promoter, the mCherry concentration does not vary with the IPTG concentration, as expected. The same experiment as in Fig 2B, but with a 10 times higher dilution rate at inoculation, giving an initial OD of 0.001. The curve shown in the plot is the same as in Fig 2B. The essential observation is that, like for an initial OD of 0.01, the switching threshold lies between 20 and 30 μM. Since for a 10 times higher dilution rate, the absorbance of growth-arrested cultures is so low that it cannot be distinguished from the background absorbance, the growth rates for IPTG concentrations between 0 and 20 μM have not been computed. Source data are available online for this figure. Download figure Download PowerPoint The results obtained with the R strain grown in minimal M9 medium with glucose suggest that for low concentrations of IPTG, the quantity of newly synthesized β and β′ subunits is not enough to sustain growth, whereas growth reaches the maximum rate for higher concentrations of IPTG. The question can be asked to which extent the observed switching behavior depends on the specific induction system used. It is well known that the response of IPTG-dependent promoters to the inducer concentration in the medium is cooperative (Kuhlman et al, 2007; Robert et al, 2010), in large part due to the positive feedback loop involved in inducer uptake. This apparent cooperativity can be strongly reduced by deleting the gene encoding the lactose transporter LacY. We therefore constructed the W-ΔlacY and R-ΔlacY strains to see whether the deletion of the lactose transporter gene affected the growth switch. As can be seen in Fig 2C, the engineered strain still functions as a growth switch and this phenotype is therefore not attributable to positive feedback in the inducer (IPTG) uptake. Does the switch also work in other growth media? In order to test this, we performed the same experiment as above in minimal M9 medium supplemented with glucose and casamino acids and in rich LB medium supplemented with glucose, both supporting higher growth rates. The results are shown in Fig 2D and E. The maximum growth rates supported by these media are indeed higher (0.016 ± 0.0005 min, corresponding to a doubling time of 42 min, in the former, and 0.027 ± 0.0008 min, corresponding to a doubling time of 27 min, in the latter). A similar growth switch is seen though, in the sense that as the inducer concentration crosses a threshold, the growth rate steeply increases from 0 to the maximum growth rate. Interestingly, as the medium becomes richer, supporting a higher maximum growth rate, the IPTG threshold at which growth starts increases as well (Fig 2F). Whereas for M9 minimal medium with glucose the switch occurs between 20 and 30 μM, supplementing this medium with casamino acids shifts the threshold to between 30 and 40 μM. In LB medium with glucose, between 40 and 50 μM of IPTG is needed for growth. The observed positive correlation between the maximum growth rate supported by a medium and the growth-switching threshold was confirmed by a fourth example, minimal M9 medium supplemented with succinate (Fig EV1). Notice that at higher growth rates proteins are diluted faster, which may require higher compensating levels of rpoBC expression. In conclusion, the results of the growth experiments demonstrate a surprising switching behavior of the strain with engineered control of rpoBC expression, occurring in several different media supporting different maximal growth rates. The control experiment with the ΔlacY strain shows that the switching behavior does not depend on the cooperativity of the classical lac induction system, but is due to internal regulatory mechanisms coupling rpoBC expression to growth. Dependence of the growth rate on the concentration of RNA polymerase While varying the IPTG concentration in the medium thus allows growth of the R strain to be" @default.
• W2175978653 created "2016-06-24" @default.
• W2175978653 creator A5022785507 @default.
• W2175978653 creator A5039330974 @default.
• W2175978653 creator A5047096952 @default.
• W2175978653 creator A5047500480 @default.
• W2175978653 creator A5054406248 @default.
• W2175978653 creator A5062022864 @default.
• W2175978653 creator A5070993495 @default.
• W2175978653 creator A5088248228 @default.
• W2175978653 creator A5091020501 @default.
• W2175978653 date "2015-11-01" @default.
• W2175978653 modified "2023-10-14" @default.
• W2175978653 title "A synthetic growth switch based on controlled expression of RNA polymerase" @default.
• W2175978653 cites W1501189928 @default.
• W2175978653 cites W1514932602 @default.
• W2175978653 cites W1561054433 @default.
• W2175978653 cites W1570124823 @default.
• W2175978653 cites W1580639168 @default.
• W2175978653 cites W1822017433 @default.
• W2175978653 cites W1966276192 @default.
• W2175978653 cites W1967104799 @default.
• W2175978653 cites W1969614792 @default.
• W2175978653 cites W1973670099 @default.
• W2175978653 cites W1977688694 @default.
• W2175978653 cites W1985161570 @default.
• W2175978653 cites W1994794118 @default.
• W2175978653 cites W1996666978 @default.
• W2175978653 cites W1998457296 @default.
• W2175978653 cites W2001357509 @default.
• W2175978653 cites W2001624473 @default.
• W2175978653 cites W2003409413 @default.
• W2175978653 cites W2005983216 @default.
• W2175978653 cites W2006718034 @default.
• W2175978653 cites W2011399510 @default.
• W2175978653 cites W2015926680 @default.
• W2175978653 cites W2023363172 @default.
• W2175978653 cites W2025105469 @default.
• W2175978653 cites W2034645234 @default.
• W2175978653 cites W2037135781 @default.
• W2175978653 cites W2037705627 @default.
• W2175978653 cites W2038173187 @default.
• W2175978653 cites W2038211892 @default.
• W2175978653 cites W2042493870 @default.
• W2175978653 cites W2044463066 @default.
• W2175978653 cites W2048299227 @default.
• W2175978653 cites W2049167322 @default.
• W2175978653 cites W2050415280 @default.
• W2175978653 cites W2059303607 @default.
• W2175978653 cites W2061659241 @default.
• W2175978653 cites W2068367947 @default.
• W2175978653 cites W2068372433 @default.
• W2175978653 cites W2068872007 @default.
• W2175978653 cites W2069759547 @default.
• W2175978653 cites W2072862994 @default.
• W2175978653 cites W2072887591 @default.
• W2175978653 cites W2076477598 @default.
• W2175978653 cites W2081781433 @default.
• W2175978653 cites W2084921586 @default.
• W2175978653 cites W2090232842 @default.
• W2175978653 cites W2092122738 @default.
• W2175978653 cites W2094117846 @default.
• W2175978653 cites W2099586831 @default.
• W2175978653 cites W2100100451 @default.
• W2175978653 cites W2100636437 @default.
• W2175978653 cites W2101762601 @default.
• W2175978653 cites W2102454474 @default.
• W2175978653 cites W2106411386 @default.
• W2175978653 cites W2107275175 @default.
• W2175978653 cites W2109242001 @default.
• W2175978653 cites W2109918485 @default.
• W2175978653 cites W2111354165 @default.
• W2175978653 cites W2111967267 @default.
• W2175978653 cites W2115606218 @default.
• W2175978653 cites W2117597548 @default.
• W2175978653 cites W2122860350 @default.
• W2175978653 cites W2123425265 @default.
• W2175978653 cites W2127223328 @default.
• W2175978653 cites W2130665506 @default.
• W2175978653 cites W2142691347 @default.
• W2175978653 cites W2144748567 @default.
• W2175978653 cites W2146500190 @default.
• W2175978653 cites W2147472054 @default.
• W2175978653 cites W2148641266 @default.
• W2175978653 cites W2152875113 @default.
• W2175978653 cites W2153326344 @default.
• W2175978653 cites W2159786000 @default.
• W2175978653 cites W2161263507 @default.
• W2175978653 cites W2161782059 @default.
• W2175978653 cites W2166374078 @default.
• W2175978653 cites W2167898143 @default.
• W2175978653 cites W2170427655 @default.
• W2175978653 cites W2301053471 @default.
• W2175978653 cites W4206619948 @default.
• W2175978653 cites W4292872153 @default.
• W2175978653 doi "https://doi.org/10.15252/msb.20156382" @default.
• W2175978653 hasPubMedCentralId "https://www.ncbi.nlm.nih.gov/pmc/articles/4670729" @default.
• W2175978653 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/26596932" @default.
• W2175978653 hasPublicationYear "2015" @default.
• W2175978653 type Work @default.

• next