Circadian control of global gene expression by the cyanobacterial master regulator RpaA
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PROVIDER: PRJNA219375 | ENA |
REPOSITORIES: ENA
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Project description:The cyanobacterium Synechococcus elongatus contains a circadian clock which coordinates circadian changes in gene expression of a large percentage of its genes. The response regulator RpaA has been implicated as an important regulator of many circadian genes, but the role of this protein in regulating changes in gene expression genome-wide is not known. We show that deletion of rpaA abrogates circadian gene expression genome-wide and arrests cells in a gene expression state highly similar to that of wildtype cells in the morning. Furthermore, we show that RpaA binds DNA in an circadian manner that is dependent on phosphorylation of the protein. To demonstrate the sufficiency of phosphorylated RpaA in driving global changes in gene expression, we used RNA sequencing to measure changes in gene expression elicited by a phosphomimetic of RpaA (RpaA D53E) and compared these changes to those that occur during a circadian cycle in wildtype cells. This analysis reveals that induction of RpaA D53E is sufficient to drive all circadian gene expression changes that happen from dawn to dusk in wildtype cells. Interestingly, the dynamics of gene expression elicited by RpaA D53E induction mirror those observed during a circadian cycle in wildtype cells, suggesting that the dynamics of circadian gene expression and hard-wired in the regulon downstream of RpaA.
2013-12-04 | GSE51112 | GEO
Project description:The cyanobacterial circadian clock generates genome-wide transcriptional oscillations and regulates cell division, but the underlying mechanisms are not well understood. Here, we show that the response regulator RpaA serves as the master regulator of these clock outputs. Deletion of rpaA abrogates gene expression rhythms globally and arrests cells in a dawn-like expression state. Although rpaA deletion causes core oscillator failure by perturbing clock gene expression, rescuing oscillator function does not restore global expression rhythms. We show that phosphorylated RpaA regulates the expression of not only clock components, generating feedback on the core oscillator, but also a small set of circadian effectors that, in turn, orchestrate genome-wide transcriptional rhythms. Expression of constitutively active RpaA is sufficient to switch cells from a dawn-like to a dusk-like expression state as well as to block cell division. Hence, complex global circadian phenotypes can be generated by controlling the phosphorylation of a single transcription factor.
| S-EPMC3935230 | biostudies-literature
Project description:The response regulator RpaA is required for control of genome-wide gene expression by the cyanobacterial circadian clock. RpaA is predicted to be a DNA binding protein based on sequence homology, but prior studies have been unable to detect binding in vitro or in vivo to a small panel of promoters. We used ChIP-Seq to determine whether RpaA associates with DNA in vivo, and if so, with what dynamics. We find that RpaA binds to over 100 location in the genome in a circadian manner, with strongest binding occuring around subjective dusk. Analysis of these binding sites shows that RpaA directly regulates the expression of clock components to generate feedback on the core oscillator, and also regulates expression of a small set of circadian effectors that in turn orchestrate global expression rhythms.
2013-12-04 | GSE51093 | GEO
Project description:The cyanobacterium Synechococcus elongatus contains a circadian clock which coordinates circadian changes in gene expression of a large percentage of its genes. The response regulator RpaA has been implicated as an important regulator of many circadian genes, but the role of this protein in regulating changes in gene expression genome-wide is not known. We show that deletion of rpaA abrogates circadian gene expression genome-wide and arrests cells in a gene expression state highly similar to that of wildtype cells in the morning. Furthermore, we show that RpaA binds DNA in an circadian manner that is dependent on phosphorylation of the protein. To demonstrate the sufficiency of phosphorylated RpaA in driving global changes in gene expression, we used RNA sequencing to measure changes in gene expression elicited by a phosphomimetic of RpaA (RpaA D53E) and compared these changes to those that occur during a circadian cycle in wildtype cells. This analysis reveals that induction of RpaA D53E is sufficient to drive all circadian gene expression changes that happen from dawn to dusk in wildtype cells. Interestingly, the dynamics of gene expression elicited by RpaA D53E induction mirror those observed during a circadian cycle in wildtype cells, suggesting that the dynamics of circadian gene expression and hard-wired in the regulon downstream of RpaA. Enriched mRNA was prepared from synchronized wildtype S. elongatus cells every four hours over a circadian cycle and sequenced using the Illumina TruSeq Stranded mRNA Sample Prep Kit and Illumina HiSeq technology. To test the role of phosphorylated RpaA in driving circadian gene expression, we generated a strain which we refer to as OX-D53E that lacks core clock components (M-NM-^TrpaA, M-NM-^TkaiBC) with an RpaA phosphomimetic (RpaA D53E) under the control of an IPTG-inducible promoter (Ptrc::rpaAD53E). We used the same methodology to measure gene expression in OX-D53E before and after induction of RpaA D53E. As a control, we also measured gene expression in the OX-D53E strain over time in the absence of IPTG. Also, we generated a strain similar to OX-D53E in which the only difference was that no gene was inserted downstream of the IPTG inducible promoter (OX-Empty). We measured gene expressio in OX-Empty before and after IPTG addition to test for off-target effects of IPTG.
2013-12-04 | E-GEOD-51112 | biostudies-arrayexpress
Project description:The response regulator RpaA is required for control of genome-wide gene expression by the cyanobacterial circadian clock. RpaA is predicted to be a DNA binding protein based on sequence homology, but prior studies have been unable to detect binding in vitro or in vivo to a small panel of promoters. We used ChIP-Seq to determine whether RpaA associates with DNA in vivo, and if so, with what dynamics. We find that RpaA binds to over 100 location in the genome in a circadian manner, with strongest binding occuring around subjective dusk. Analysis of these binding sites shows that RpaA directly regulates the expression of clock components to generate feedback on the core oscillator, and also regulates expression of a small set of circadian effectors that in turn orchestrate global expression rhythms. Crosslinked samples were acquired every four hours from a turbidostatic wild-type (AMC408) cultures in constant light (LL) following entrainment with two light-dark (LD) cycles. As a negative control, we acquired samples similarly from an M-NM-^TrpaA culture. Chromatin immunoprecipitation was performed on each sample from wht wild-type and from a pool of all samples from the M-NM-^TrpaA culture. Libraries were prepared from the ChIP samples and sequenced with Illumina technology. For the wild-type, biological replicate samples were acquired at times of maximum and minimum RpaA binding.
2013-12-04 | E-GEOD-51093 | biostudies-arrayexpress
Project description:This SuperSeries is composed of the SubSeries listed below. Refer to individual Series
2013-12-04 | E-GEOD-50922 | biostudies-arrayexpress
Project description:This SuperSeries is composed of the SubSeries listed below.
2013-12-04 | GSE50922 | GEO
Project description:The transcription factor RpaA is the master regulator of circadian transcription in cyanobacteria, driving genome-wide oscillations in mRNA abundance. Deletion of rpaA has no effect on viability in constant light conditions, but renders cells inviable in cycling conditions when light and dark periods alternate. We investigated the mechanisms underlying this viability defect. We performed RNA-seq experiment using the rpaA- “clock rescue” and “clock rescue” strains and we show that the rpaA- “clock rescue” strain is defective in transcription of genes crucial for utilization of carbohydrate stores at night.
2017-02-15 | GSE89999 | GEO
Project description:Like many other organisms, cyanobacteria exhibit rhythmic gene expression with a period length of 24 hours to adapt to daily environmental changes. In the model organism Synechococcus elongatus PCC 7942 the central oscillator consists of three proteins: KaiA, KaiB and KaiC and utilizes the histidine kinase SasA and its response regulator RpaA as output-signaling pathway. Synechocystis sp. PCC 6803 contains two additional homologs of the kaiB and kaiC genes. Here we demonstrate that RpaA interacts with the core oscillator KaiAB1C1 of Synechocystis sp. PCC 6803 via SasA, similar to Synechococcus elongatus PCC 7942. However, interaction with the additional Kai homologs was not detected, suggesting different signal transduction components for the clock homologs. Inactivation of rpaA in Synechocystis sp. PCC 6803, lead to reduced viability of the mutant in light-dark cycles that aggravated under mixotrophic growth conditions. Chemoheterotrophic growth in the dark was abolished completely. In accordance, transcriptomic data revealed that RpaA is involved in the regulation of genes related to CO2‑acclimation and carbon metabolism under diurnal light conditions. Further, our results indicate that RpaA functions in the posttranslational regulation of glycogen metabolism as well, and a potential link between the circadian clock and motility was identified.
2018-10-01 | GSE116795 | GEO
Project description:Zhou2015 - Circadian clock with immune
regulator NPR1
Arabidopsis clock model modified from
P2012 (Pokhilko et al., 2013 -
BIOMD0000000445)
model to include the master immune regulator NPR1 coupling to LHY,
TOC1 and PRR7.
Triggers: The Global Quantities contain triggers that allow
one to change coupling settings, Salicyclic acid (SA) treatment and
npr1 mutants.
LHY_on: true->NPR1 couples to LHY
PRR7_on: true->NPR1 couples to PRR7
WT: true->WT plants, false->npr1 mutant plants
SA: true->SA treated plants, false->no treatment
This model has L=1, i.e. operates only under constant light
conditions and is not aiming to make preditions under diurnal
conditions. Due to period overshoot only time points after 28h are
relevant.
This model is described in the article:
Redox rhythm reinforces the
circadian clock to gate immune response.
Zhou M, Wang W, Karapetyan S, Mwimba
M, Marqués J, Buchler NE, Dong X.
Nature 2015 Jun;
Abstract:
Recent studies have shown that in addition to the
transcriptional circadian clock, many organisms, including
Arabidopsis, have a circadian redox rhythm driven by the
organism's metabolic activities. It has been hypothesized that
the redox rhythm is linked to the circadian clock, but the
mechanism and the biological significance of this link have
only begun to be investigated. Here we report that the master
immune regulator NPR1 (non-expressor of pathogenesis-related
gene 1) of Arabidopsis is a sensor of the plant's redox state
and regulates transcription of core circadian clock genes even
in the absence of pathogen challenge. Surprisingly, acute
perturbation in the redox status triggered by the immune signal
salicylic acid does not compromise the circadian clock but
rather leads to its reinforcement. Mathematical modelling and
subsequent experiments show that NPR1 reinforces the circadian
clock without changing the period by regulating both the
morning and the evening clock genes. This balanced network
architecture helps plants gate their immune responses towards
the morning and minimize costs on growth at night. Our study
demonstrates how a sensitive redox rhythm interacts with a
robust circadian clock to ensure proper responsiveness to
environmental stimuli without compromising fitness of the
organism.
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2024-09-02 | BIOMD0000000577 | BioModels