Project description:To investigate acclimation mechanisms employed under extreme high light conditions, gene expression analysis was performed using the model microalgae Synechocystis sp. PCC 6803 (PCC 6803) cultured under various light intensities. From the low to the mid light conditions, the expression of genes related to light harvesting systems was repressed, whereas that of CO2 fixation and of D1 protein turnover-related genes was induced. Gene expression data also revealed that the down-regulation of genes related to flagellum synthesis (pilA2), pyridine nucleotide transhydrogenase (pntA and pntB), and sigma factor (sigA and sigF) represents acclimation mechanisms of PCC 6803 under excessive high light conditions.
Project description:In cyanobacteria DNA supercoiling varies over the diurnal light/dark cycle and is integrated with temporal programs of transcription and replication. We manipulated DNA supercoiling in Synechocystis sp. PCC 6803 by CRISPRi-based knockdown of gyrase subunits gyrA, gyrB and overexpression of topoisomerase I (TopoI) topA and analyzed the transcriptional response to gyrase knock-downs (endpoint in triplicate) and topoisomerase I overexpression (endpoint in triplicate, and 19 time points time series before and after induction) in Synechocystis sp. PCC 6803 via RNA-seq of coding RNA. In detail, Illumina Ribo-Zero Plus rRNA Depletion Kit was used to remove the ribosomal RNA molecules from the isolated total RNA. Removal of rRNA was evaluated with the RNA Pico 6000 kit on the Agilent 2100 Bioanalyzer. RNA was free of detectable rRNA. Preparation of cDNA libraries was performed according to the manufacturer’s instructions for the TruSeq stranded mRNA kit (Illumina, San Diego, CA, United States). Subsequently, each cDNA library was sequenced on an Illumina NextSeq 500 system (2 x 75 nt PE high output v2.5).
Project description:we obtained a HL tolerant (Tol) strain Synechocystis sp. PCC 6803 by adaptive evolution experiment that the cells were repeatedly subcultured for prolonged periods of time (52 days) under high light stress condition (7000 to 9000 μmol m-2 s-1). Although the growth of the parental strain almost stopped under 9000 μmol m-2 s-1, no growth inhibition was observed in the Tol strain. Furthermore, the growth rate was identical to that of parental strain under low light condition (40 μmol m-2 s-1). To further investigate the high light tolerant mechanisms in the Tol strain, the transcriptome was performed. The transcriptome data suggests the increase of isiA expression in the Tol strain under HL condition. The overexpression of isiA successfully enhanced the HL stress tolerance in the parental strain. The HL tolerant mechanism was different from previous reported mechanisms, such as a reduction of the light-harvesting antenna size. The tolerant strain would be an attractive host for bio-production under high light conditions.
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.
