Mesophyll conductance response to short-term changes in pCO2 is related to leaf anatomy and biochemistry in diverse C4 grasses

Mesophyll CO2 conductance (gm) in C3 species responds to short-term (minutes) changes in environment potentially due to changes in some leaf anatomical and biochemical properties and due to measurement artifacts. Compared to C3 species, there is less information about gm responses to short-term changes in environmental conditions like pCO2 across diverse C4 species and the potential determinants of these responses. Using 16 diverse C4-grasses we investigated the response of gm to short-term changes in pCO2 and how this response related to the leaf anatomical and biochemical traits. For all the measured C4-grasses gm increased as CO2 decreased; however, the percent change in gm varied (+13% to +250%) and significantly related to percent changes in leaf transpiration efficiency (TEi). The percent increase in gm was highest in grasses with thinner mesophyll cell walls and greater leaf nitrogen, activities of phosphoenolpyruvate carboxylase (PEPC), Rubisco, and carbonic anhydrase, and a higher affinity of PEPC for bicarbonate substrate. Our study demonstrates that CO2 response of gm varies greatly across diverse C4-grasses and identifies the key leaf anatomical and biochemical traits related to this variation. These findings have implications for improving C4 photosynthetic models, and in attempts to improve TEi through manipulation of gm.

Comparative analysis morphology, anatomical structure and transcriptional regulatory network of chlorophyll biosynthesis in Oryza longistaminata, O. sativa and their F1 generation

Oryza longistaminata, a perennial wild species, is widely distributed in the African continent. It has strong tolerance to biotic and abiotic stresses, and high biomass production on poor soils. Chlorophyll biosynthesis is important for photosynthesis in rice. However, the chlorophyll biosynthesis and related gene profiles of O. longistaminata and its descendants remained unclear. Here, the F1 generation of O. sativa and O. longistaminata were obtained. Then, the comparative analysis morphology, anatomical structure, and transcriptional regulatory networks of chlorophyll biosynthesis were detected and analyzed. Results showed that the F1 generation has obvious long awn, similar with that of the male parent. The purple color of the long awn is different from that of the male parent. Microstructural results showed that the flag leaves of F1 have large mesophyll cell gaps in the upper- and lower-positions, small mesophyll cell gaps in the middle position, and more chloroplasts. Increased chlorophyll content was also observed in the F1 generation. In the lower-position flag leaves, the total chlorophyll contents of F1 were 1.55 and 1.5 times those of O. sativa and O. longistaminata, respectively. POR, MgCH and HEMA1 showed higher expression levels than the other related genes selected in the chlorophyll biosynthesis pathway. The HEMA1 expression level in the middle-position flag leaves of O. longistaminata was the highest, and it was 2.83 and 2.51 times that of O. sativa and F1, respectively. The expression level of DVR gene in lower-position flag leaves of F1 were 93.16% and 95.06% lower than those of O. sativa and O. longistaminata, respectively. This study provided a potential reference for studying the photosynthesis and heterosis utilization of O. longistaminata.

Physiological And Proteomic Analyses Reveals That Brassinosteroids Application Improves The Chilling Stress Tolerance of Pepper Seedlings

Brassinosteroids (BRs) are important in plant resistance to chilling stress. However, limited information is available regarding the specific mechanisms involved at proteomic level. We utilized iTRAQ proteomic approach, physiological assays and information obtained from cellular ultrastructure to clarify the underlying molecular mechanism of BRs to alleviate chilling stress in pepper (Capsicum annuum L.). Foliar application of 24-epibrassinolide (EBR) improved photosynthesis and improved cell structure by presenting a distinct mesophyll cell and chloroplast with well-developed thylakoid membranes in the leaves of pepper seedlings. We identified 346 differentially expressed proteins (DEPs), including 217 up-regulated proteins and 129 down-regulated proteins in plants under chilling (Chill) and Chill + EBR treated plants. Most of the DEPs were related to multiple pathways, including photosynthesis, carbohydrates metabolism, energy metabolism, protein biosynthesis, amino acids synthesis, redox and stress defence (ascorbate peroxidase, glutathione peroxidase, and superoxide dismutase). Up-regulated DEPs were associated with photosynthetic electron transfer chain, oxidative phosphorylation, GSH metabolism pathway, Calvin cycle and signaling pathway. The physiochemical analysis showed that EBR treatment improved the tolerance of pepper seedlings to chilling stress.

CO2-Responsive CCT Protein interacts with 14-3-3 proteins and controls the expression of starch synthesis-related genes

CO2 responsive CCT protein (CRCT) is a positive regulator of starch synthesis related genes such as ADP-glucose pyrophosphorylase large subunit 1 and starch branching enzyme I particularly in the leaf sheath of rice (Oryza sativa L.). The promoter GUS analysis revealed that CRCT expressed exclusively in the vascular bundle, whereas starch synthesis related genes were expressed in different sites such as mesophyll cell and starch storage parenchyma cell. However, the chromatin immunoprecipitation (ChIP) using a FLAG-CRCT overexpression line and subsequent qPCR analyses showed that the 5’-flanking regions of these starch synthesis-related genes tended to be enriched by ChIP, suggesting that CRCT can bind to the promoter regions of these genes. The monomer of CRCT is 34.2 kDa, however CRCT was detected at 270 kDa via gel filtration chromatography, suggesting that CRCT forms a complex in vivo. Immunoprecipitation and subsequent MS analysis pulled down several 14-3-3-like proteins. A yeast two-hybrid analysis and bimolecular fluorescence complementation assays confirmed the interaction between CRCT and 14-3-3-like proteins. Although there is an inconsistency in the place of expression, this study provide important findings regarding the molecular function of CRCT to control the expression of key starch synthesis-related genes.

CO 2 diffusion in tobacco: a link between mesophyll conductance and leaf anatomy

The partial pressure of CO 2 at the sites of carboxylation within chloroplasts depends on the conductance to CO 2 diffusion from intercellular airspace to the sites of carboxylation, termed mesophyll conductance ( g m ). We investigated how g m varies with leaf age and through a tobacco ( Nicotiana tabacum ) canopy by combining gas exchange and carbon isotope measurements using tunable diode laser spectroscopy. We combined these measurements with the anatomical characterization of leaves. CO 2 assimilation rate, A , and g m decreased as leaves aged and moved lower in the canopy and were linearly correlated. This was accompanied by large anatomical changes including an increase in leaf thickness. Chloroplast surface area exposed to the intercellular airspace per unit leaf area ( S c ) also decreased lower in the canopy. Older leaves had thicker mesophyll cell walls and g m was inversely proportional to cell wall thickness. We conclude that reduced g m of older leaves lower in the canopy was associated with a reduction in S c and a thickening of mesophyll cell walls.

Effect of lead treatment on the stipe and leaflet anatomy of Pteris vittata and Pityrogramma calomelanos (Pteridaceae: fern)

The effect of various concentrations of Lead on stipe and leaflet anatomy of Pteris vittata and Pityrogramma calomelanos were investigated. Fernlets were transplanted into 5 kg pots of soil pre-treated with five different concentrations of Lead, adequately watered and arranged in a complete randomized design in the screen house. Pots with no Lead treatment served as control. At twelve weeks after planting, plants were carefully removed, washed and taken to the laboratory. Anatomical sections of leaflets and stipe of each of the treatments were carried out with a sledge microtome (10-15 μm). Microscopic observations of structures were made after staining the sections. In Pteris vittata, the thickness of the cuticle of the stipe increased with increasing Lead concentrations but the reverse of this was observed in Pityrogamma calomelanos. There was also significant reduction of epidermal and mesophyll cell area of both species with increasing Lead concentrations. It can be concluded from this study that P. vittata has a higher tolerance level to Lead than Pityrogramma calomelanos. Key words: Anatomy, Cuticle thickness, Ferns, Lead, Pteridaceae.

Plant resistance in different cell layers affects aphid probing and feeding behaviour during non-host and poor-host interactions

Aphids are phloem-feeding insects that cause economic losses to crops globally. Whilst aphid interactions with susceptible plants and partially resistant genotypes have been well characterized, the interactions between aphids and non-host species are not well understood. Unravelling these non-host interactions can identify the mechanisms which contribute to plant resistance. Using contrasting aphid-host plant systems, including the broad host range pest Myzus persicae (host: Arabidopsis; poor-host: barley) and the cereal pest Rhopalosiphum padi (host: barley; non-host: Arabidopsis), we conducted a range of physiological experiments and compared aphid settling and probing behaviour on a host plant vs either a non-host or poor-host. In choice experiments, we observed that around 10% of aphids selected a non-host or poor-host plant species after 24 h. Using the Electrical Penetration Graph technique, we showed that feeding and probing behaviours differ during non-host and poor-host interactions when compared with a host interaction. In the Arabidopsis non-host interaction with the cereal pest R. padi aphids were unable to reach and feed on the phloem, with resistance likely residing in the mesophyll cell layer. In the barley poor-host interaction with M. persicae, resistance is likely phloem-based as phloem ingestion was reduced compared with the host interaction. Overall, our data suggest that plant resistance to aphids in non-host and poor-host interactions with these aphid species likely resides in different plant cell layers. Future work will take into account specific cell layers where resistances are based to dissect the underlying mechanisms and gain a better understanding of how we may improve crop resistance to aphids.

Variability in the chloroplast area lining the intercellular airspace and cell walls drives mesophyll conductance in gymnosperms

The photosynthetic efficiency of plants in different environments is controlled by stomata, hydraulics, biochemistry, and mesophyll conductance (gm). Recently, gm was demonstrated to be the key limitation of photosynthesis in gymnosperms. Values of gm across gymnosperms varied over 20-fold, but this variation was poorly explained by robust structure-bound integrated traits such as leaf dry mass per area. Understanding how the component structural traits control gm is central for identifying the determinants of variability in gm across plant functional and phylogenetic groups. Here, we investigated the structural traits responsible for gm in 65 diverse gymnosperms. Although the integrated morphological traits, shape, and anatomical characteristics varied widely across species, the distinguishing features of all gymnosperms were thick mesophyll cell walls and low chloroplast area exposed to intercellular airspace (Sc/S) compared with angiosperms. Sc/S and cell wall thickness were the fundamental traits driving variations in gm across gymnosperm species. Chloroplast thickness was the strongest limitation of gm among liquid-phase components. The variation in leaf dry mass per area was not correlated with the key ultrastructural traits determining gm. Thus, given the absence of correlating integrated easy-to-measure traits, detailed knowledge of underlying component traits controlling gm across plant taxa is necessary to understand the photosynthetic limitations across ecosystems.