Two KaiABC systems control circadian oscillations in one cyanobacterium.

The circadian clock of cyanobacteria, which predicts daily environmental changes, typically includes a standard oscillator consisting of proteins KaiA, KaiB, and KaiC. However, several cyanobacteria have diverse Kai protein homologs of unclear function. In particular, Synechocystis sp.

Reduced expression of chlorophyllide oxygenase (CAO) decreases the metabolic flux for chlorophyll synthesis and downregulates photosynthesis in tobacco plants.

Unlabelled: Chlorophyll is synthesized from chlorophyllide , catalyzed by chlorophyllide oxygenase (). To examine whether reduced chlorophyll content regulates chlorophyll (Chl) synthesis and photosynthesis, we raised transgenic tobacco plants with antisense expression, which had lower chlorophyll content and, thus, higher Chl / ratio. Further, these plants had (i) lower chlorophyll and total Chl content, whether they were grown under low or high light; (ii) decreased steady-state levels of chlorophyll biosynthetic intermediates, due, perhaps, to a feedback-controlled reduction in enzyme expressions/activities; (iii) reduced electron transport rates in their intact leaves, and reduced Photosystem (PS) I, PS II and whole chain electron transport activities in their isolated thylakoids; (iv) decreased carbon assimilation in plants grown under low or high light.

KidA, a multi-PAS domain protein, tunes the period of the cyanobacterial circadian oscillator.

The cyanobacterial clock presents a unique opportunity to understand the biochemical basis of circadian rhythms. The core oscillator, composed of the KaiA, KaiB, and KaiC proteins, has been extensively studied, but a complete picture of its connection to the physiology of the cell is lacking. To identify previously unknown components of the clock, we used KaiB locked in its active fold as bait in an immunoprecipitation/mass spectrometry approach.

Daily Cycles of Reversible Protein Condensation in Cyanobacteria.

An emerging principle of cell biology is the regulated conversion of macromolecules between soluble and condensed states. To screen for such regulation of the cyanobacterial proteome, we use quantitative mass spectrometry to identify proteins that change solubility during the day-night cycle. We find a set of night-insoluble proteins that includes many enzymes in essential metabolic pathways.

Kalman-like Self-Tuned Sensitivity in Biophysical Sensing.

Living organisms need to be sensitive to a changing environment while also ignoring uninformative environmental fluctuations. Here, we argue that living cells can navigate these conflicting demands by dynamically tuning their environmental sensitivity. We analyze the circadian clock in Synechococcus elongatus, showing that clock-metabolism coupling can detect mismatch between clock predictions and the day-night light cycle, temporarily raise the clock's sensitivity to light changes, and thus re-entraining faster.

Modulation of biosynthesis of photosynthetic pigments and light-harvesting complex in wild-type and gun5 mutant of Arabidopsis thaliana during impaired chloroplast development.

Plants in response to different environmental cues need to modulate the expression of nuclear and chloroplast genomes that are in constant communication. To understand the signals that are responsible for inter-organellar communication, levulinic acid (LA), an inhibitor of 5-aminolevulinic acid dehydratase, was used to suppress the synthesis of pyrrole-derived tetrapyrroles chlorophylls. Although, it does not specifically inhibit carotenoid biosynthesis enzymes, LA reduced the carotenoid contents during photomorphogenesis of etiolated Arabidopsis seedlings.

Controlling the Cyanobacterial Clock by Synthetically Rewiring Metabolism.

Circadian clocks are oscillatory systems that allow organisms to anticipate rhythmic changes in the environment. Several studies have shown that circadian clocks are connected to metabolism, but it is not generally clear whether metabolic signaling is one voice among many that influence the clock or whether metabolic cycling is the major clock synchronizer. To address this question in cyanobacteria, we used a synthetic biology approach to make normally autotrophic cells capable of growth on exogenous sugar.

Rhythms in energy storage control the ability of the cyanobacterial circadian clock to reset.

Circadian clocks are oscillatory systems that schedule daily rhythms of organismal behavior. The ability of the clock to reset its phase in response to external signals is critical for proper synchronization with the environment. In the model clock from cyanobacteria, the KaiABC proteins that comprise the core oscillator are directly sensitive to metabolites.

The cyanobacterial clock and metabolism.

Cyanobacteria possess the simplest known circadian clock, which presents a unique opportunity to study how rhythms are generated and how input signals from the environment reset the clock time. The kaiABC locus forms the core of the oscillator, and the remarkable ability to reconstitute oscillations using purified KaiABC proteins has allowed researchers to study mechanism using the tools of quantitative biochemistry. Autotrophic cyanobacteria experience major shifts in metabolism following a light-dark transition, and recent work suggests that input mechanisms that couple the day-night cycle to the clock involve energy and redox metabolites acting directly on clock proteins.

Light intensity-dependent modulation of chlorophyll b biosynthesis and photosynthesis by overexpression of chlorophyllide a oxygenase in tobacco.

Chlorophyll b is synthesized by the oxidation of a methyl group on the B ring of a tetrapyrrole molecule to a formyl group by chlorophyllide a oxygenase (CAO). The full-length CAO from Arabidopsis (Arabidopsis thaliana) was overexpressed in tobacco (Nicotiana tabacum) that grows well at light intensities much higher than those tolerated by Arabidopsis. This resulted in an increased synthesis of glutamate semialdehyde, 5-aminolevulinic acid, magnesium-porphyrins, and chlorophylls.

Overexpression of protochlorophyllide oxidoreductase C regulates oxidative stress in Arabidopsis.

Light absorbed by colored intermediates of chlorophyll biosynthesis is not utilized in photosynthesis; instead, it is transferred to molecular oxygen, generating singlet oxygen ((1)O(2)). As there is no enzymatic detoxification mechanism available in plants to destroy (1)O(2), its generation should be minimized. We manipulated the concentration of a major chlorophyll biosynthetic intermediate i.

Accelerated cell death 2 suppresses mitochondrial oxidative bursts and modulates cell death in Arabidopsis.

The Arabidopsis ACCELERATED CELL DEATH 2 (ACD2) protein protects cells from programmed cell death (PCD) caused by endogenous porphyrin-related molecules like red chlorophyll catabolite or exogenous protoporphyrin IX. We previously found that during bacterial infection, ACD2, a chlorophyll breakdown enzyme, localizes to both chloroplasts and mitochondria in leaves. Additionally, acd2 cells show mitochondrial dysfunction.

Light-dependent regulation of chlorophyll b biosynthesis in chlorophyllide a oxygenase overexpressing tobacco plants.

Chlorophyllide a oxygenase (CAO) that converts chlorophyllide a to chlorophyllide b was overexpressed in tobacco to increase chlorophyll (Chl) b biosynthesis and alter the Chl a/b ratio. Transgenic plants along with their wild-type cultivars were grown in low and high light intensities. In low light there was 20% increase in chlorophyll b contents in transgenic plants, which resulted in 16% reduction in the Chl a/b ratio.

Catalytic function of a novel protein protochlorophyllide oxidoreductase C of Arabidopsis thaliana.

In Arabidopsis thaliana Por C has been identified only on sequence homology to that of por A and por B. To demonstrate its catalytic function Arabidopsis thaliana protochlorophyllide oxidoreductase C gene (por c) that codes for the mature part of POR C protein having 335 amino acids was expressed in Escherchia coli cells. The POR C enzyme in the presence of NADPH and protochlorophyllide when incubated in dark formed a ternary complex.