scholarly journals ldpA Encodes an Iron-Sulfur Protein Involved in Light-Dependent Modulation of the Circadian Period in the Cyanobacterium Synechococcuselongatus PCC 7942

2003 ◽  
Vol 185 (4) ◽  
pp. 1415-1422 ◽  
Author(s):  
Mitsunori Katayama ◽  
Takao Kondo ◽  
Jin Xiong ◽  
Susan S. Golden
Keyword(s):  
Phase Shift ◽  
Circadian Clock ◽  
Binding Motif ◽  
Circadian Period ◽  
Wild Type ◽  
Reading Frame ◽  
Short Period ◽  
Screening Protocol ◽  
Dark Pulse ◽  
Pcc 7942

ABSTRACT We generated random transposon insertion mutants to identify genes involved in light input pathways to the circadian clock of the cyanobacterium Synechococcus elongatus PCC 7942. Two mutants, AMC408-M1 and AMC408-M2, were isolated that responded to a 5-h dark pulse differently from the wild-type strain. The two mutants carried independent transposon insertions in an open reading frame here named ldpA (for light-dependent period). Although the mutants were isolated by a phase shift screening protocol, the actual defect is a conditional alteration in the circadian period. The mutants retain the wild-type ability to phase shift the circadian gene expression (bioluminescent reporter) rhythm if the timing of administration of the dark pulse is corrected for a 1-h shortening of the circadian period in the mutant. Further analysis indicated that the conditional short-period mutant phenotype results from insensitivity to light gradients that normally modulate the circadian period in S. elongatus, lengthening the period at low light intensities. The ldpA gene encodes a polypeptide that predicts a 7Fe-8S cluster-binding motif expected to be involved in redox reactions. We suggest that the LdpA protein modulates the circadian clock as an indirect function of light intensity by sensing changes in cellular physiology.

scholarly journals Rhizosphere microbes influence host circadian clock function

2018 ◽  
Author(s):  
Charley J. Hubbard ◽  
Robby McMinn ◽  
Cynthia Weinig
Keyword(s):  
Circadian Clock ◽  
Microbial Communities ◽  
Circadian Period ◽  
Wild Type ◽  
Host Genotype ◽  
Local Conditions ◽  
Mutant Genotype ◽  
Individual Fitness ◽  
Short Period ◽  
Clock Mutant

AbstractThe circadian clock is an important determinant of individual fitness that is entrained by local conditions. In addition to known abiotic inputs that entrain the circadian clock, individual pathogenic soil bacteria affect the circadian period of plant hosts. Yet, in nature, plants interact with diverse microbial communities including hundreds to thousands of microbial taxa, and the effect of these communities on clock function remains unclear. In Arabidopsis thaliana, we used diverse rhizosphere inoculates and both wild-type and clock mutant genotypes to test the effect of complex rhizosphere microbial communities on the host circadian clock. Host plants with an intact rhizosphere microbiome expressed a circadian period that was closer to 24 hrs in duration and significantly shorter (by 60 minutes on average) relative to plants grown with a disrupted microbiome. Wild-type host genotypes differed significantly in clock sensitivity to microbiome treatments, where the effect was most pronounced in the Landsberg erecta genotype and least in the Columbia genotype. Rhizosphere microbes collected from a host genotype with a short-period phenotype (toc1-21) and used as inoculate significantly shortened the long-period phenotype of the ztl-1 clock mutant genotype. The results indicate that complex rhizosphere microbial communities significantly affect host clock function.

scholarly journals A Novel Mutation in kaiC Affects Resetting of the Cyanobacterial Circadian Clock

2005 ◽  
Vol 187 (8) ◽  
pp. 2559-2564 ◽  
Author(s):  
Yota B. Kiyohara ◽  
Mitsunori Katayama ◽  
Takao Kondo
Keyword(s):  
Circadian Rhythm ◽  
Circadian Rhythms ◽  
Phase Shift ◽  
Feedback Inhibition ◽  
Novel Mutation ◽  
Feedback Regulation ◽  
Phase Shifting ◽  
Wild Type ◽  
Dark Pulse ◽  
Pcc 7942

ABSTRACT Light is the most important factor controlling circadian systems in response to day-night cycles. In order to better understand the regulation of circadian rhythms by light in Synechococcus elongatus PCC 7942, we screened for mutants with defective phase shifting in response to dark pulses. Using a 5-h dark-pulse protocol, we identified a mutation in kaiC that we termed pr1, for phase response 1. In the pr1 mutant, a 5-h dark pulse failed to shift the phase of the circadian rhythm, while the same pulse caused a 10-h phase shift in wild-type cells. The rhythm in accumulation of KaiC was abolished in the pr1 mutant, and the rhythmicity of KaiC phosphorylation was reduced. Additionally, the pr1 mutant was defective in mediating the feedback inhibition of kaiBC. Finally, overexpression of mutant KaiC led to a reduced phase shift compared to that for wild-type KaiC. Thus, KaiC appears to play a role in resetting the cellular clock in addition to its documented role in the feedback regulation of circadian rhythms.

scholarly journals The Synechococcus Strain PCC 7942glnN Product (Glutamine Synthetase III) Helps Recovery from Prolonged Nitrogen Chlorosis

2000 ◽  
Vol 182 (19) ◽  
pp. 5615-5619 ◽  
Author(s):  
Jörg Sauer ◽  
Ulrike Dirmeier ◽  
Karl Forchhammer
Keyword(s):  
Glutamine Synthetase ◽  
Transcriptional Start Site ◽  
Binding Motif ◽  
Class Iii ◽  
Wild Type ◽  
Cloning And Sequencing ◽  
Gene Encoding ◽  
Low Concentrations ◽  
Synechococcus Strain ◽  
Pcc 7942

ABSTRACT We report the cloning and sequencing of the glnN gene encoding a class III glutamine synthetase from the cyanobacteriumSynechococcus strain PCC 7942. Mapping of the transcriptional start site revealed a DNA sequence in the promoter region that resembles an imperfect NtcA binding motif. Expression ofglnN is impaired in NtcA- and PII-deficient mutants. The only parameter which was negatively affected in theglnN mutant compared to the wild type was the recovery rate of prolonged nitrogen-starved cells with low concentrations of combined nitrogen.

scholarly journals Rhizosphere microbes influence host circadian clock function

2021 ◽  
Author(s):  
Charley Hubbard ◽  
Robby McMinn ◽  
Cynthia Weinig
Keyword(s):  
Arabidopsis Thaliana ◽  
Circadian Clock ◽  
Microbial Communities ◽  
Abiotic Factors ◽  
Circadian Period ◽  
Local Conditions ◽  
Mutant Genotype ◽  
Short Period ◽  
Clock Mutant ◽  
Boechera Stricta

The circadian clock is an important determinant of fitness that is entrained by local conditions. Aside from abiotic factors, individual pathogenic soil bacteria affect circadian clock function in plant hosts. Yet, in nature, plants interact with diverse microbial communities, and the effect of complex communities on clock function remains unclear. In Arabidopsis thaliana and its wild relative, Boechera stricta, we used diverse rhizosphere inoculates and host genotypes to test the effect of complex rhizosphere microbial communities on the host circadian clock. Arabidopsis thaliana plants with an intact rhizosphere microbiome expressed a circadian period closer to 24h in duration and significantly shorter (by 48 minutes on average) relative to plants grown with a disrupted microbiome. Wild-type host genotypes of A. thaliana differed in clock sensitivity to microbes, with one genotype (Landsberg erecta) expressing a 119-minute difference in circadian period length across rhizosphere microbial treatments. A similar pattern of clock sensitivity to soil microbes was observed in B. stricta. Finally, rhizosphere microbes collected from the mutant genotype toc1-21 of A. thaliana with a short-period phenotype and used as inoculate significantly shortened the long-period phenotype of the clock mutant genotype ztl-1. The results indicate that complex rhizosphere microbial communities affect host clock function.

scholarly journals fleN, a Gene That Regulates Flagellar Number in Pseudomonas aeruginosa

2000 ◽  
Vol 182 (2) ◽  
pp. 357-364 ◽  
Author(s):  
Nandini Dasgupta ◽  
Shiwani K. Arora ◽  
Reuben Ramphal
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Binding Motif ◽  
Gene Promoters ◽  
Wild Type ◽  
Electron Microscopic ◽  
Genome Database ◽  
Reading Frame ◽  
Mutant Strains ◽  
In The Wild ◽  
Flagellar Genes

ABSTRACT The single polar flagellum of Pseudomonas aeruginosaplays an important role in the pathogenesis of infection by this organism. However, regulation of the assembly of this organelle has not been delineated. In analyzing the sequence available at thePseudomonas genome database, an open reading frame (ORF), flanked by flagellar genes flhF and fliA, that coded for a protein (280 amino acids) with an ATP-binding motif at its N terminus was found. The ORF was inactivated by inserting a gentamicin cassette in P. aeruginosa PAK and PAO1. The resulting mutants were nonmotile on motility agar plates, but under a light microscope they exhibited random movement and tumbling behavior. Electron microscopic studies of the wild-type and mutant strains revealed that the mutants were multiflagellate, with three to six polar flagella per bacterium as rather than one as in the wild type, indicating that this ORF was involved in regulating the number of flagella and chemotactic motility in P. aeruginosa. The ORF was named fleN. An intact copy of fleN on a plasmid complemented the mutant by restoring motility and monoflagellate status. The β-galactosidase activities of eight flagellar operon or gene promoters in the wild-type andfleN mutant strains revealed a direct correlation between six promoters that were upregulated in the fleN mutant (fliLMNOPQ, flgBCDE, fliEFG,fliDS orf126, fleSR, and fliC) and positive regulation by FleQ, an NtrC-like transcriptional regulator for flagellar genes. Based on these results, we propose a model where FleN influences FleQ activity (directly or indirectly) in regulating flagellar number in P. aeruginosa.

scholarly journals Changing planetary rotation rescues the biological clock mutantlhy cca1ofArabidopsis thaliana

2015 ◽  
Author(s):  
Andrew J. Millar ◽  
Jamie T. Carrington ◽  
Wei Ven Tee ◽  
Sarah K. Hodge
Keyword(s):  
Gene Expression ◽  
Circadian Clock ◽  
Double Mutant ◽  
Clock Genes ◽  
Biological Clock ◽  
Laboratory Model ◽  
Wild Type ◽  
Planetary Rotation ◽  
Short Period ◽  
Clock Mechanism

Background: Pervasive, 24-hour rhythms from the biological clock affect diverse biological processes in metabolism and behaviour, including the human cell division cycle and sleep-wake cycle, nightly transpiration and energy balance in plants, and seasonal breeding in both plants and animals. The clock mechanism in the laboratory model plant species Arabidopsis thaliana is complex, in part due to the multiple interlocking, negative feedback loops that link the clock genes. Clock gene mutants are powerful tools to manipulate and understand the clock mechanism and its effects on physiology. The LATE ELONGATED HYPOCOTYL and CIRCADIAN CLOCK ASSOCIATED 1 genes encode dawn-expressed, Myb-related repressor proteins that delay the expression of other clock genes until late in the day. Double mutant plants (lhy cca1) have low-amplitude, short-period rhythms that have been used in multiple studies of the plant circadian clock. Results: We used in vivo imaging of several luciferase (LUC) reporter genes to test how the rhythmic gene expression of wild-type and lhy cca1 mutant plants responded to light:dark cycles. Red, blue and red+blue light were similarly able to entrain these gene expression rhythms. The timing of expression rhythms in double mutant plants showed little or no response to the duration of light under 24h light:dark cycles (dusk sensitivity), in contrast to the wild type. As the period of the mutant clock is about 18h, we tested light:dark cycles of different duration (T cycles), simulating altered rotation of planet Earth. lhy cca1 double mutants regained as much dusk sensitivity in 20h T cycles as the wild type in 24h cycles, though the phase of the rhythm in the mutants was much earlier than wild type. The severe, triple lhy cca1 gi mutants also regained dusk sensitivity in 20h cycles. The double mutant showed some dusk sensitivity under 28h cycles. lhy cca1 double mutants under 28h cycles with short photoperiods, however, had the same apparent phase as wild-type plants. Conclusion: Simulating altered planetary rotation with light:dark cycles can reveal normal circadian performance in clock mutants that have been described as arrhythmic under standard conditions. The features rescued here comprise a dynamic behaviour (apparent phase under 28h cycles) and a dynamic property (dusk sensitivity under 20h cycles). These conditional clock phenotypes indicate that parts of the clock mechanism continue to function independently of LHY and CCA1, despite the major role of these genes in wild-type plants under standard conditions. Accessibility: Most results here will be published only in this format, citable by the DOI. Data and analysis are publicly accessible on the BioDare resource (www.biodare.ed.ac.uk), as detailed in the links below. Transgenic lines are linked to Stock Centre IDs below (Table 7).

scholarly journals The Circadian Clock-Related Gene pex Regulates a Negative cis Element in the kaiA Promoter Region

2007 ◽  
Vol 189 (21) ◽  
pp. 7690-7696 ◽  
Author(s):  
Shinsuke Kutsuna ◽  
Takao Kondo ◽  
Haruki Ikegami ◽  
Tatsuya Uzumaki ◽  
Mitsunori Katayama ◽  
...  
Keyword(s):  
Circadian Clock ◽  
Constitutive Expression ◽  
Negative Regulation ◽  
Start Codon ◽  
Upstream Region ◽  
Related Gene ◽  
Wild Type ◽  
In The Wild ◽  
Expression Activity ◽  
Pcc 7942

ABSTRACT In the cyanobacterium Synechococcus sp. strain PCC 7942, a circadian clock-related gene, pex, was identified as the gene prolonging the period of the clock. A PadR domain, which is a newly classified transcription factor domain, and the X-ray crystal structure of the Pex protein suggest a role for Pex in transcriptional regulation in the circadian system. However, the regulatory target of the Pex protein is unknown. To determine the role of Pex, we monitored bioluminescence rhythms that reported the expression activity of the kaiA gene or the kaiBC operon in pex deficiency, pex constitutive expression, and the wild-type genotype. The expression of kaiA in the pex-deficient or constitutive expression genotype was 7 or 1/7 times that of the wild type, respectively, suggesting that kaiA is the target of negative regulation by Pex. In contrast, the expression of the kaiBC gene in the two pex-related genotypes was the same as that in the wild type, suggesting that Pex specifically regulates kaiA expression. We used primer extension analysis to map the transcription start site for the kaiA gene 66 bp upstream of the translation start codon. Mapping with deletion and base pair substitution of the kaiA upstream region revealed that a 5-bp sequence in this region was essential for the regulation of kaiA. The repression or constitutive expression of the kaiA transgene caused the prolongation or shortening of the circadian period, respectively, suggesting that the Pex protein changes the period via the negative regulation of kaiA.

The short-period mutant, toc1-1, alters circadian clock regulation of multiple outputs throughout development in Arabidopsis thaliana

Development ◽  
1998 ◽  
Vol 125 (3) ◽  
pp. 485-494 ◽  
Author(s):  
D.E. Somers ◽  
A.A. Webb ◽  
M. Pearson ◽  
S.A. Kay
Keyword(s):  
Arabidopsis Thaliana ◽  
Circadian Clock ◽  
Red Light ◽  
Luciferase Reporter ◽  
Wild Type ◽  
Luciferase Reporter Gene ◽  
Environmental Signals ◽  
Developmental States ◽  
Short Period ◽  
Log Linear

The coordination of developmental and physiological events with environmental signals is facilitated by the action of the circadian clock. Here we report a new set of circadian clock-controlled phenotypes for Arabidopsis thaliana. We use these markers together with the short-period mutant, toc1-1, and the clock-controlled cab2::luciferase reporter gene to assess the nature of the circadian clock throughout development and to suggest the position of TOC1 within the circadian clock system. In dark-grown seedlings, the toc1-1 lesion conferred a short period to the cycling of cab2::luciferase luminescence, as previously found in light-grown plants, indicating that the circadian clocks in these two divergent developmental states share at least one component. Stomatal conductance rhythms were similarly approximately 3 hours shorter than wild type in toc1-1, suggesting that a cell-autonomous clockwork may be active in guard cells in 5- to 6-week-old leaves. The effect of daylength on flowering time in the C24 ecotype was diminished by toc1-1, and was nearly eliminated in the Landsberg erecta background where the plants flowered equally early in both short and long days. Throughout a 500-fold range of red light intensities, both the wild type and the mutant showed an inverse log-linear relationship of fluence rate to period, with a 2–3 hour shorter period for the mutant at all intensities. These results indicate that TOC1 acts on or within the clock independently of light input. Temperature entrainment appears normal in toc1-1, and the period-shortening effects of the mutant remain unchanged over a 20 degrees C temperature range. Taken together our results are consistent with the likelihood that TOC1 codes for an oscillator component rather than for an element of an input signaling pathway. In addition, the pervasive effect of toc1-1 on a variety of clock-controlled processes throughout development suggests that a single circadian system is primarily responsible for controlling most, if not all, circadian rhythms in the plant.

scholarly journals A Novel Allele of kaiA Shortens the Circadian Period and Strengthens Interaction of Oscillator Components in the Cyanobacterium Synechococcus elongatus PCC 7942

2009 ◽  
Vol 191 (13) ◽  
pp. 4392-4400 ◽  
Author(s):  
You Chen ◽  
Yong-Ick Kim ◽  
Shannon R. Mackey ◽  
C. Kay Holtman ◽  
Andy LiWang ◽  
...  
Keyword(s):  
Protein Complexes ◽  
Model Organism ◽  
Synechococcus Elongatus ◽  
Luciferase Reporter ◽  
Rhythmic Pattern ◽  
Circadian Period ◽  
C Terminus ◽  
Short Period ◽  
Synechococcus Elongatus Pcc 7942 ◽  
Pcc 7942

ABSTRACT The basic circadian oscillator of the unicellular fresh water cyanobacterium Synechococcus elongatus PCC 7942, the model organism for cyanobacterial circadian clocks, consists of only three protein components: KaiA, KaiB, and KaiC. These proteins, all of which are homomultimers, periodically interact to form large protein complexes with stoichiometries that depend on the phosphorylation state of KaiC. KaiA stimulates KaiC autophosphorylation through direct physical interactions. Screening a library of S. elongatus transposon mutants for circadian clock phenotypes uncovered an atypical short-period mutant that carries a kaiA insertion. Genetic and biochemical analyses showed that the short-period phenotype is caused by the truncation of KaiA by three amino acid residues at its C terminus. The disruption of a negative element upstream of the kaiBC promoter was another consequence of the insertion of the transposon; when not associated with a truncated kaiA allele, this mutation extended the circadian period. The circadian rhythm of KaiC phosphorylation was conserved in these mutants, but with some modifications in the rhythmic pattern of KaiC phosphorylation, such as the ratio of phosphorylated to unphosphorylated KaiC and the relative phase of the circadian phosphorylation peak. The results showed that there is no correlation between the phasing of the KaiC phosphorylation pattern and the rhythm of gene expression, measured as bioluminescence from luciferase reporter genes. The interaction between KaiC and the truncated KaiA was stronger than normal, as shown by fluorescence anisotropy analysis. Our data suggest that the KaiA-KaiC interaction and the circadian pattern of KaiC autophosphorylation are both important for determining the period, but not the relative phasing, of circadian rhythms in S. elongatus.

scholarly journals The Cloning and Molecular Analysis of pawn-B in Paramecium tetraurelia

Genetics ◽  
2000 ◽  
Vol 155 (3) ◽  
pp. 1105-1117 ◽  
Author(s):  
W John Haynes ◽  
Kit-Yin Ling ◽  
Robin R Preston ◽  
Yoshiro Saimi ◽  
Ching Kung
Keyword(s):  
Genomic Library ◽  
Open Reading Frame ◽  
Paramecium Tetraurelia ◽  
Wild Type ◽  
Rt Pcr ◽  
Reading Frame ◽  
Close Proximity ◽  
In The Wild ◽  
Dna Fragment ◽  
Alternative Sites

Abstract Pawn mutants of Paramecium tetraurelia lack a depolarization-activated Ca2+ current and do not swim backward. Using the method of microinjection and sorting a genomic library, we have cloned a DNA fragment that complements pawn-B (pwB/pwB). The minimal complementing fragment is a 798-bp open reading frame (ORF) that restores the Ca2+ current and the backward swimming when expressed. This ORF contains a 29-bp intron and is transcribed and translated. The translated product has two putative transmembrane domains but no clear matches in current databases. Mutations in the available pwB alleles were found within this ORF. The d4-95 and d4-96 alleles are single base substitutions, while d4-662 (previously pawn-D) harbors a 44-bp insertion that matches an internal eliminated sequence (IES) found in the wild-type germline DNA except for a single C-to-T transition. Northern hybridizations and RT-PCR indicate that d4-662 transcripts are rapidly degraded or not produced. A second 155-bp IES in the wild-type germline ORF excises at two alternative sites spanning three asparagine codons. The pwB ORF appears to be separated from a 5′ neighboring ORF by only 36 bp. The close proximity of the two ORFs and the location of the pwB protein as indicated by GFP-fusion constructs are discussed.

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