ABSTRACT
Fructose-1,6-bisphosphatase (FBPase) and sedoheptulose-1,7-bisphosphatase (SBPase) are two essential activities in the Calvin-Benson-Bassham cycle that catalyze two irreversible reactions and are key for proper regulation and functioning of the cycle. These two activities are codified by a single gene in all cyanobacteria, although some cyanobacteria contain an additional gene coding for a FBPase. Mutants lacking the gene coding for SBP/FBPase protein are not able to grow photoautotrophically and require glucose to survive. As this protein presents both activities, we have tried to elucidate which of the two are required for photoautrophic growth in Synechocystis sp PCC 6803. For this, the genes coding for plant FBPase and SBPase were introduced in a SBP/FBPase mutant strain, and the strains were tested for growth in the absence of glucose. Ectopic expression of only a plant SBPase gene did not allow growth in the absence of glucose although allowed mutation of both Synechocystis' FBPase genes. When both plant FBPase and SBPase genes were expressed, photoautrophic growth of the SBP/FBPase mutants was restored. This complementation was partial as the strain only grew in low light, but growth was impaired at higher light intensities. Redox regulation of the Calvin-Benson-Bassham cycle is essential to properly coordinate light reactions to carbon fixation in the chloroplast. Two of the best characterized proteins that are redox-regulated in the cycle are FBPase and SBPase. These two proteins are targets of the FTR-Trx redox system with Trx f being the main reductant in vivo. Introduction of the TrxF gene improves growth of the complemented strain, suggesting that the redox state of the proteins may be the cause of this phenotype. The redox state of the plant proteins was also checked in these strains, and it shows that the cyanobacterial redox system is able to reduce all of them (SBPase, FBPase, and TrxF) in a light-dependent manner. Thus, the TrxF-FBPase-SBPase plant chloroplast system is active in cyanobacteria despite that these organisms do not contain proteins related to them. Furthermore, our system opens the possibility to study specificity of the Trx system in vivo without the complication of the different isoforms present in plants.
ABSTRACT
After the Great Oxidation Event (GOE), iron availability was greatly decreased, and photosynthetic organisms evolved several alternative proteins and mechanisms. One of these proteins, plastocyanin, is a type I blue-copper protein that can replace cytochrome c6 as a soluble electron carrier between cytochrome b6f and photosystem I. In most cyanobacteria, expression of these two alternative proteins is regulated by copper availability, but the regulatory system remains unknown. Herein, we provide evidence that the regulatory system is composed of a BlaI/CopY-family transcription factor (PetR) and a BlaR-membrane protease (PetP). PetR represses petE (plastocyanin) expression and activates petJ (cytochrome c6), while PetP controls PetR levels in vivo. Using whole-cell extracts, we demonstrated that PetR degradation requires both PetP and copper. Transcriptomic analysis revealed that the PetRP system regulates only four genes (petE, petJ, slr0601, and slr0602), highlighting its specificity. Furthermore, the presence of petE and petRP in early branching cyanobacteria indicates that acquisition of these genes could represent an early adaptation to decreased iron bioavailability following the GOE.