SCARECROW gene function is required for photosynthetic development in maize

C4 photosynthesis in grasses relies on a specialized leaf anatomy. In maize, this ‘Kranz’ leaf anatomy is patterned in part by the duplicated SCARECROW (SCR) genes ZmSCR1 and ZmSCR1h. Here we show that in addition to patterning defects, chlorophyll content and levels of transcripts encoding Golden2-like regulators of chloroplast development are significantly lower in Zmscr1;Zmscr1h mutants than in wild-type. These perturbations are not associated with changes in chloroplast number, size or ultrastructure. However, the maximum rates of carboxylation by ribulose bisphosphate carboxylase/oxygenase (RuBisCO, Vcmax) and phosphoenolpyruvate carboxylase (PEPC, Vpmax) are both reduced, leading to perturbed plant growth. The CO2 compensation point and 13C‰ of Zmscr1;Zmscr1h plants are both normal, indicating that a canonical C4 cycle is operating, albeit at reduced overall capacity. Taken together, our results reveal that the maize SCR genes, either directly or indirectly, play a role in photosynthetic development.Significance statementSCARECROW (SCR) is one of the best studied plant developmental regulators, however, its role in downstream plant physiology is less well-understood. Here, we have demonstrated that SCR is required to establish and/or maintain photosynthetic capacity in maize leaves.

SlSNAT Interacts with HSP40, a Molecular Chaperone, to Regulate Melatonin Biosynthesis and Promote Thermotolerance in Tomato

The SNAT enzyme participates in the biosynthesis of melatonin, which is reported to regulate thermotolerance in many plants. However, the mechanistic basis of this regulation remains unclear. In this study, we identified the SlSNAT gene, which is responsible for melatonin biosynthesis in tomato. SlSNAT expression levels were 3- and 5-fold higher in SlSNAT overexpression lines OX-2 and OX-6, respectively. The melatonin levels were 3- and 4-fold higher than those in wild type. The melatonin levels decreased by 50% when the expression of SlSNAT was downregulated to 40%. Overexpression of SlSNAT in tomato plants provided significantly enhanced thermotolerance with better growth performance in Photosystem II (PSII) maximum photochemical quantum yield (Fv/Fm) and alleviated heat injury. Both exogenous melatonin treatment and endogenous melatonin manipulation by SlSNAT overexpression decreased the levels of reactive oxygen species�accumulation and Fv/Fm. The SlSNAT overexpression line showed protected ribulose bisphosphate carboxylase oxygenase proteins and upregulated response of heat transcription factors and heat shock proteins under heat stress. HSP40, a DnaJ-type chaperone, was found to interact with SlSNAT in the chloroplast. Downregulation of HSP40 showed lower melatonin synthesis under heat stress. HSP40 functions as a chaperone to protect the SNAT enzyme during melatonin synthesis under heat stress. HSP40 interacted with SlSNAT and together participated in melatonin-related thermotolerance regulation in tomato.

Proteomic analyses of Citrus petiole responses to early Huanglongbing Disease

Backgroud Huanglongbing (HLB) is currently one of the most destructive citrus disease worldwide. It is caused by Candidatus Liberibacter asiaticus (CLas), a nonculturable alpha-proteobacterium, which it resides exclusively in the phloem tissues. Therefore, understanding the early CLas-responsive proteins in citrus petiole where pathogenic bacteria colonized will help to investigate plant resistance to the pathogen.Results In this study, a comparative proteomic approach was applied to identify the petiole proteins associated with the response to CLas infection. A total of 777 proteins were differentially expressed in response to CLas. Among them, 499 proteins were up-regulated and 278 were down-regulated. Among the most highly up-regulated differentially expressed proteins (DEPs) were salicylate carboxymethyltransferase, ubiquitin carboxyl-terminal hydrolase 13, trans-resveratrol di-O-methyltransferase, linoleate 13S-lipoxygenase 2-1, granule-bound starch synthase 1 and thaumatin-like proteins. While the most highly down-regulated DEPs were oxygen-evolving enhancer proteins, ribulose bisphosphate carboxylase/oxygenase activase, peroxidases and photosystem reaction center subunits. The results of qPCR analysis of a number of indicated DEPs and western blotting further validated four representative DEPs, including salicylate carboxymethyltransferase, linoleate 13S-lipoxygenase 2-1, granule-bound starch synthase 1 and photosystem I reaction center subunit showed that most of detected DEPs were positively correlated with their mRNA and protein levels.Conclusions Our comparative proteomic analysis first profiling reveals early and primary proteome alterations in CLas-infected citrus petiole, where pathogens reside in. The DEPs results demonstrate that CLas infection could promote the carbohydrate metabolism, depress the photosystem and activate/inhibit defense responses.

Form III RubisCO-mediated transaldolase variant of the Calvin cycle in a chemolithoautotrophic bacterium

The Calvin–Benson–Bassham (CBB) cycle assimilates CO2for the primary production of organic matter in all plants and algae, as well as in some autotrophic bacteria. The key enzyme of the CBB cycle, ribulose-bisphosphate carboxylase/oxygenase (RubisCO), is a main determinant of de novo organic matter production on Earth. Of the three carboxylating forms of RubisCO, forms I and II participate in autotrophy, and form III so far has been associated only with nucleotide and nucleoside metabolism. Here, we report that form III RubisCO functions in the CBB cycle in the thermophilic chemolithoautotrophic bacteriumThermodesulfobium acidiphilum,a phylum-level lineage representative. We further show that autotrophic CO2fixation inT. acidiphilumis accomplished via the transaldolase variant of the CBB cycle, which has not been previously demonstrated experimentally and has been considered unlikely to occur. Thus, this work reveals a distinct form of the key pathway of CO2fixation.

Thylakoid localized bestrophin-like proteins are essential for the CO2 concentrating mechanism of Chlamydomonas reinhardtii

The green alga Chlamydomonas reinhardtii possesses a CO2 concentrating mechanism (CCM) that helps in successful acclimation to low CO2 conditions. Current models of the CCM postulate that a series of ion transporters bring HCO3− from outside the cell to the thylakoid lumen, where the carbonic anhydrase 3 (CAH3) dehydrates accumulated HCO3− to CO2, raising the CO2 concentration for Ribulose bisphosphate carboxylase/oxygenase (Rubisco). Previously, HCO3− transporters have been identified at both the plasma membrane and the chloroplast envelope, but the transporter thought to be on the thylakoid membrane has not been identified. Three paralogous genes (BST1, BST2, and BST3) belonging to the bestrophin family have been found to be up-regulated in low CO2 conditions, and their expression is controlled by CIA5, a transcription factor that controls many CCM genes. YFP fusions demonstrate that all 3 proteins are located on the thylakoid membrane, and interactome studies indicate that they might associate with chloroplast CCM components. A single mutant defective in BST3 has near-normal growth on low CO2, indicating that the 3 bestrophin-like proteins may have redundant functions. Therefore, an RNA interference (RNAi) approach was adopted to reduce the expression of all 3 genes at once. RNAi mutants with reduced expression of BST1–3 were unable to grow at low CO2 concentrations, exhibited a reduced affinity to inorganic carbon (Ci) compared with the wild-type cells, and showed reduced Ci uptake. We propose that these bestrophin-like proteins are essential components of the CCM that deliver HCO3− accumulated in the chloroplast stroma to CAH3 inside the thylakoid lumen.

pH determines the energetic efficiency of the cyanobacterial CO2 concentrating mechanism

Many carbon-fixing bacteria rely on a CO2 concentrating mechanism (CCM) to elevate the CO2 concentration around the carboxylating enzyme ribulose bisphosphate carboxylase/oxygenase (RuBisCO). The CCM is postulated to simultaneously enhance the rate of carboxylation and minimize oxygenation, a competitive reaction with O2 also catalyzed by RuBisCO. To achieve this effect, the CCM combines two features: active transport of inorganic carbon into the cell and colocalization of carbonic anhydrase and RuBisCO inside proteinaceous microcompartments called carboxysomes. Understanding the significance of the various CCM components requires reconciling biochemical intuition with a quantitative description of the system. To this end, we have developed a mathematical model of the CCM to analyze its energetic costs and the inherent intertwining of physiology and pH. We find that intracellular pH greatly affects the cost of inorganic carbon accumulation. At low pH the inorganic carbon pool contains more of the highly cell-permeable H2CO3, necessitating a substantial expenditure of energy on transport to maintain internal inorganic carbon levels. An intracellular pH ≈8 reduces leakage, making the CCM significantly more energetically efficient. This pH prediction coincides well with our measurement of intracellular pH in a model cyanobacterium. We also demonstrate that CO2 retention in the carboxysome is necessary, whereas selective uptake of HCO3− into the carboxysome would not appreciably enhance energetic efficiency. Altogether, integration of pH produces a model that is quantitatively consistent with cyanobacterial physiology, emphasizing that pH cannot be neglected when describing biological systems interacting with inorganic carbon pools.

Algal evolution in relation to atmospheric CO 2 : carboxylases, carbon-concentrating mechanisms and carbon oxidation cycles

Oxygenic photosynthesis evolved at least 2.4 Ga; all oxygenic organisms use the ribulose bisphosphate carboxylase-oxygenase (Rubisco)–photosynthetic carbon reduction cycle (PCRC) rather than one of the five other known pathways of autotrophic CO 2 assimilation. The high CO 2 and (initially) O 2 -free conditions permitted the use of a Rubisco with a high maximum specific reaction rate. As CO 2 decreased and O 2 increased, Rubisco oxygenase activity increased and 2-phosphoglycolate was produced, with the evolution of pathways recycling this inhibitory product to sugar phosphates. Changed atmospheric composition also selected for Rubiscos with higher CO 2 affinity and CO 2 /O 2 selectivity correlated with decreased CO 2 -saturated catalytic capacity and/or for CO 2 -concentrating mechanisms (CCMs). These changes increase the energy, nitrogen, phosphorus, iron, zinc and manganese cost of producing and operating Rubisco–PCRC, while biosphere oxygenation decreased the availability of nitrogen, phosphorus and iron. The majority of algae today have CCMs; the timing of their origins is unclear. If CCMs evolved in a low-CO 2 episode followed by one or more lengthy high-CO 2 episodes, CCM retention could involve a combination of environmental factors known to favour CCM retention in extant organisms that also occur in a warmer high-CO 2 ocean. More investigations, including studies of genetic adaptation, are needed.

Atomic Resolution X-ray Structure of the Substrate Recognition Domain of Higher Plant Ribulose-bisphosphate Carboxylase/Oxygenase (Rubisco) Activase

The rapid release of tight-binding inhibitors from dead-end ribulose-bisphosphate carboxylase/oxygenase (Rubisco) complexes requires the activity of Rubisco activase, an AAA+ ATPase that utilizes chemo-mechanical energy to catalyze the reactivation of Rubisco. Activase is thought to play a central role in coordinating the rate of CO2 fixation with the light reactions of photosynthesis. Here, we present a 1.9 Å crystal structure of the C-domain core of creosote activase. The fold consists of a canonical four-helix bundle, from which a paddle-like extension protrudes that entails a nine-turn helix lined by an irregularly structured peptide strand. The residues Lys-313 and Val-316 involved in the species-specific recognition of Rubisco are located near the tip of the paddle. An ionic bond between Lys-313 and Glu-309 appears to stabilize the glycine-rich end of the helix. Structural superpositions onto the distant homolog FtsH imply that the paddles extend away from the hexameric toroid in a fan-like fashion, such that the hydrophobic sides of each blade bearing Trp-302 are facing inward and the polar sides bearing Lys-313 and Val-316 are facing outward. Therefore, we speculate that upon binding, the activase paddles embrace the Rubisco cylinder by placing their hydrophobic patches near the partner protein. This model suggests that conformational adjustments at the remote end of the paddle may relate to selectivity in recognition, rather than specific ionic contacts involving Lys-313. Additionally, the superpositions predict that the catalytically critical Arg-293 does not interact with the bound nucleotide. Hypothetical ring-ring stacking and peptide threading models for Rubisco reactivation are briefly discussed.