Cyanobacteriochromes are a newly recognized group of photoreceptors that are distinct relatives of phytochromes but are found only in cyanobacteria. A putative cyanobacteriochrome, CcaS, is known to chromatically regulate the expression of the phycobilisome linker gene (cpcG2) in Synechocystis sp. PCC 6803. In this study, we isolated the chromophore-binding domain of CcaS from Synechocystis as well as from phycocyanobilin-producing Escherichia coli. Both preparations showed the same reversible photoconversion between a green-absorbing form (Pg, max ‫؍‬ 535 nm) and a red-absorbing form (Pr, max ‫؍‬ 672 nm). Mass spectrometry and denaturation analyses suggested that Pg and Pr bind phycocyanobilin in a double-bond configuration of C15-Z and C15-E, respectively. Autophosphorylation activity of the histidine kinase domain in nearly full-length CcaS was up-regulated by preirradiation with green light. Similarly, phosphotransfer to the cognate response regulator, CcaR, was higher in Pr than in Pg. From these results, we conclude that CcaS phosphorylates CcaR under green light and induces expression of cpcG2, leading to accumulation of CpcG2-phycobilisome as a chromatic acclimation system. CcaS is the first recognized green light receptor in the expanded phytochrome superfamily, which includes phytochromes and cyanobacteriochromes.chromatic adaptation ͉ phycocyanobilin ͉ phytochrome ͉ cyanobacteria ͉ photoreceptor P hytochromes (Phys) are photoreceptors that typically perceive red and far-red light and regulate a wide range of physiological responses in plants, bacteria, cyanobacteria, and fungi (1). They exhibit reversible photoconversion between two distinct forms: the red-absorbing form (Pr) and the far-redabsorbing form (Pfr). Their N-terminal photosensory region, which consists of Per-ARNT-Sim (PAS), cGMP phosphodiesterase/adenylyl cyclase/FhlA (GAF), and phytochrome domains, is highly conserved, but there are variations in the chromophore of the linear tetrapyrrole, such as phytochromobilin, phycocyanobilin (PCB), and biliverdin. It is reported that phytochromobilin or PCB is covalently anchored at a conserved cysteine residue in the GAF domain (2, 3), whereas biliverdin is anchored at another conserved cysteine residue in the N terminus of the PAS domain (4). The perception of light by Phys triggers a Z to E isomerization of the C15-C16 double bond between the C and D pyrrole rings as well as subsequent conformational changes of the chromophore and the apoprotein [supporting information (SI) Fig. S1] (5) which signal to downstream processes. Recent crystallographic analyses of bacterial Phys (DrBphP and RpBphP3) have revealed the three-dimensional structure of PAS and GAF domains in the Pr form (6-8). The biliverdin chromophore is buried deep within a pocket in the GAF domain with a configuration of C5-Z,syn/C10-Z,syn/C15-Z,anti. Because the residues in the chromophore-binding pocket are highly conserved, it was proposed that Phys share a common photoconversion mechanism, albeit with certain variations.''Cyanobac...

The circadian oscillator of the cyanobacterium Synechococcus elongatus, like those in eukaryotes, is entrained by environmental cues. Inactivation of the gene cikA (circadian input kinase) shortens the circadian period of gene expression rhythms in S. elongatus by approximately 2 hours, changes the phasing of a subset of rhythms, and nearly abolishes resetting of phase by a pulse of darkness. The CikA protein sequence reveals that it is a divergent bacteriophytochrome with characteristic histidine protein kinase motifs and a cryptic response regulator motif. CikA is likely a key component of a pathway that provides environmental input to the circadian oscillator in S. elongatus.

;The gene, pixJ1 (formerly pisJ1), is predicted to encode a phytochrome-like photoreceptor that is essential for positive phototaxis in the unicellular cyanobacterium

BLUF (a sensor of Blue-Light Using FAD) is a novel putative photoreceptor domain that is found in many bacteria and some eukaryotic algae. As found on genome analysis, certain cyanobacteria have BLUF proteins with a short C-terminal extension. As typical examples, Tll0078 from thermophilic Thermosynechococcus elongatus BP-1 and Slr1694 from mesophilic Synechocystis sp. PCC 6803 were comparatively studied. FAD of both proteins was hardly reduced by exogenous reductants or mediators except methylviologen but showed a typical spectral shift to a longer wavelength upon excitation with blue light. In particular, freshly prepared Tll0078 protein showed slow but reversible aggregation, indicative of light-induced conformational changes in the protein structure. Tll0078 is far more stable as to heat treatment than Slr1694, as judged from flavin fluorescence. The slr1694-disruptant showed phototactic motility away from the light source (negative phototaxis), while the wild type Synechocystis showed positive phototaxis toward the source. Yeast two-hybrid screening with slr1694 showed self-interaction of Slr1694 (PixD) with itself and interaction with a novel PatA-like response regulator, Slr1693 (PixE). These results were discussed in relation to the signaling mechanism of the "short" BLUF proteins in the regulation of cyanobacterial phototaxis.

Cyanobacteria have several putative photoreceptors (designated cyanobacteriochromes) that are related to but distinct from the established phytochromes. The GAF domain of the phototaxis regulator, PixJ, from a thermophilic cyanobacterium Thermosynechococcus elongatus BP-1 (TePixJ_GAF) is a cyanobacteriochrome which exhibits reversible photoconversion between a blue light-absorbing form (max = 433 nm) and a green light-absorbing form (max = 531 nm). To study the chromophore, we prepared TePixJ_GAF chromoprotein from heterologously expressed Synechocystis and performed spectral analysis after denaturation by comparing it with the cyanobacterial phytochrome Cph1 which harbors phycocyanobilin (PCB) as a chromophore. The results indicated that the chromophore of TePixJ is not PCB, but its isomer, phycoviolobilin (PVB). It is suggested that the GAF domain of TePixJ has auto-lyase and auto-isomerase activities.

A kaiABC clock gene cluster was previously identified from cyanobacterium Synechococcus elongatus PCC 7942, and the feedback regulation of kai genes was proposed as the core mechanism generating circadian oscillation. In this study, we confirmed that the Kai-based oscillator is the dominant circadian oscillator functioning in cyanobacteria. We probed the nature of this regulation and found that excess KaiC represses not only kaiBC but also the rhythmic components of all genes in the genome. This result strongly suggests that the KaiC protein primarily coordinates genomewide gene expression, including its own expression. We also found that a promoter derived from E. coli is feedback controlled by KaiC and restores the complete circadian rhythm in kaiBC-inactivated arrhythmic mutants, provided it can express kaiB and kaiC genes at an appropriate level. Unlike eukaryotic models, specific regulation of the kaiBC promoter is not essential for cyanobacterial circadian oscillations.

Responding to green and red light, certain cyanobacteria change the composition of their light-harvesting pigments, phycoerythrin (PE) and phycocyanin (PC). Although this phenomenon—complementary chromatic adaptation—is well known, the green light–sensing mechanism for PE accumulation is unclear. The filamentous cyanobacterium Nostoc punctiforme ATCC 29133 ( N. punctiforme ) regulates PE synthesis in response to green and red light (group II chromatic adaptation). We disrupted the green/red-perceiving histidine-kinase gene ( ccaS ) or the cognate response regulator gene ( ccaR ), which are clustered with several PE and PC genes ( cpeC - cpcG2-cpeR1 operon) in N. punctiforme . Under green light, wild-type cells accumulated a significant amount of PE upon induction of cpeC - cpcG2 - cpeR1 expression, whereas they accumulated little PE with suppression of cpeC - cpcG2 - cpeR1 expression under red light. Under both green and red light, the ccaS mutant constitutively accumulated some PE with constitutively low cpeC - cpcG2 - cpeR1 expression, whereas the ccaR mutant accumulated little PE with suppression of cpeC - cpcG2 - cpeR1 expression. The results of an electrophoretic mobility shift assay suggest that CcaR binds to the promoter region of cpeC - cpcG2 - cpeR1 , which contains a conserved direct-repeat motif. Taken together, the results suggest that CcaS phosphorylates CcaR under green light and that phosphorylated CcaR then induces cpeC - cpcG2 - cpeR1 expression, leading to PE accumulation. In contrast, CcaS probably represses cpeC - cpcG2 - cpeR1 expression by dephosphorylation of CcaR under red light. We also found that the cpeB-cpeA operon is partially regulated by green and red light, suggesting that the green light-induced regulatory protein CpeR1 activates cpeB-cpeA expression together with constitutively induced CpeR2.

In the cyanobacterium Synechococcus elongatus PCC 7942, circadian timing is transmitted from the KaiABC-based central oscillator to the transcription factor RpaA via the KaiC-interacting histidine kinase SasA to activate transcription, thereby generating rhythmic circadian gene expression. However, KaiC can also repress circadian gene expression, including its own. The mechanism and significance of this negative feedback regulation have been unclear. Here, we report a novel gene, labA (low-amplitude and bright), that is required for negative feedback regulation of KaiC. Disruption of labA abolished transcriptional repression caused by overexpression of KaiC and elevated the trough levels of circadian gene expression, resulting in a low-amplitude phenotype. In contrast, overexpression of labA significantly lowered circadian gene expression. Furthermore, genetic analysis indicated that labA and sasA function in parallel pathways to regulate kaiBC expression, whereas rpaA functions downstream from labA for kaiBC expression. These results suggest that temporal information from the KaiABC-based oscillator diverges into a LabA-dependent negative pathway and a SasA-dependent positive pathway, and then converges onto RpaA to generate robust circadian gene expression. It is likely that quantitative information of KaiC is transmitted to RpaA through LabA, whereas SasA mediates the state of the KaiABC-based oscillator.[Keywords: Circadian clock; cyanobacteria; KaiC; labA; RpaA; SasA] Supplemental material is available at http://www.genesdev.org. The circadian clock is an endogenous timing system that controls various biological activities with a period of ∼24 h. Most organisms use self-sustained oscillation to coordinate with and adapt to daily environmental changes. KaiA, KaiB, and KaiC have been identified as essential components for circadian oscillation in the cyanobacterium Synechococcus elongatus PCC 7942 (hereafter Synechococcus). KaiC, an autokinase and autophosphatase, is the central component of the cyanobacterial circadian clock and interacts with KaiA and KaiB Iwasaki et al. 1999;Nishiwaki et al. 2000;Taniguchi et al. 2001). KaiA enhances the autokinase activity of KaiC (Iwasaki et al. 2002;Williams et al. 2002;Uzumaki et al. 2004) and/or inhibits its autophosphatase activity (Kitayama et al. 2003;Xu et al. 2003), while KaiB attenuates the activity of KaiA (Williams et al. 2002;Kitayama et al. 2003;Xu et al. 2003). KaiC phosphorylation oscillates with a period of ∼24 h when the three recombinant Kai proteins are incubated in vitro in the presence of ATP . This oscillation period is refractory to changes in temperature, an important feature of circadian rhythms ). Thus, Kai-based chemical oscillation is thought to be the basic circadian timing loop in Synechococcus Tomita et al. 2005).KaiC also interacts with a sensory histidine kinase, SasA . Autophosphorylation of SasA is enhanced in response to KaiC binding (Smith and Williams 2006;Takai et al. 2006), and this phosphate group is then transferred to the putati...