Whole-genome gene expression profiling in the cyanobacterium Synechococcus sp. PCC 7002 : impacts of electron transfer mutations and low CO₂ stress
Cyanobacteria are the ancestors of plant chloroplasts and perform ~25% of global photosynthesis. The photosynthetic reaction center complexes (PSII and PSI) capture solar energy to catalyze a series of electron transfer, oxidation/reduction (redox) reactions along the pathway: H2O!PSII!plastoquinone (PQ) pool!cytochrome (Cyt) bf complex!PSI!ferredoxin. These reactions generate a transmembrane proton gradient for adenosine triphosphate (ATP) synthesis and reduce nicotine adenine dinucleotide (NADP+) to generate NAD(P)H. ATP and NAD(P)H provide the energy and electrons to drive carbon fixation, the conversion of CO2 into carbon polymers. The Cyt bf complex is implicated in sensing the redox potential of the PQ pool and signaling adaptive changes in photosynthesis and gene expression. This thesis research tested the hypothesis that the Cyt bf complex mediates redox-dependent signaling of gene expression and sought to identify the gene targets of this regulation. Specifically, mutants defective in Cyt bf lowand high-potential electron transfer chains were used to investigate the role of these pathways in redox signaling during optimal photosynthesis and low CO2 stress. Custom high-density oligonucleotide microarrays (NimbleGen®) and commercial (DNAStar®), as well as open-source bioinformatics programs (Bioconductor R) were used to investigate whole-genome gene expression responses and identify transcription start sites of 94 key photosynthesis genes in the cyanobacterium Synechococcus sp. PCC 7002. These microarray studies identified ~250 genes (~13% of the genome) differentially upregulated genes. Transcription start site mapping revealed that 87 of 94 genes had upstream transcriptional activity. 52 of 94 genes had transcripts beginning within ~100-300 base pairs of their known start codons. 34 of 94 genes showed continuous transcription with neighboring genes, indicating operon structures. Specifically, this work identified an operon of carbon assimilation genes (rbcR–ndhF3–ndhD3–cupA–A0175) upregulated under low CO2 as in the cyanobacterium Synechocystis sp. PCC 6803. In contrast to the wild type, Cyt bf low- and high-potential chain mutants (PetB-R214H and PetC1-!2G, respectively) upregulated a bicarbonate Na+/H+ antiporter (bicA) operon under low CO2, suggesting the induction of an additional carbon acquisition response in these mutants. Uniquely, the Cyt bf low-potential chain mutant upregulated a petF gene for ferredoxin, the electron acceptor for PSI. The Cyt bf high-potential chain mutant upregulated ndhD for a subunit of the NAD(P)H dehydrogenase (NDH-1) complex. Both PSI and NDH-1 are important for adjusting the redox balance of electron transfer and photosynthesis. These altered gene expression patterns in the Cyt bf low- and high potential chain mutants provide evidence that the distribution of electron flow in the Cyt bf complex plays a central role in regulating redox potential and photosynthesis. This research contributes to understanding redox regulation of gene expression in cyanobacteria andto strategies for bioproduct-biofuels production.