The relationship between contents of photosynthetic metabolites and the rate of photosynthetic carbon assimilation in leaves of Amaranthus edulis L.

The relationship between the gas-exchange characteristics of attached leaves of Amaranthus edulis L. and the contents of photosynthetic intermediates was examined in response to changing irradiance and intercellular partial pressure of CO₂. After determination of the rate of CO₂ assimilation at known intercellular CO₂ pressure and irradiance, the leaf was freeze-clamped and the contents of ribulose-1,5-bisphosphate, glycerate-3-phosphate, fructose-1,6-bisphosphate, glucose-6-phosphate, fructose-6-phosphate, triose phosphates, phosphoenolpyruvate, pyruvate, oxaloacetate, aspartate, alanine, malate and glutamate were measured. A comparison between the sizes of metabolite pools and theoretical calculations of metabolite gradients required for transport between the mesophyll and the bundle-sheath cells showed that aspartate, alanine, glycerate-3-phosphate and triose phosphates were present in sufficient quantities to support transport by diffusion, whereas pyruvate and oxaloacetate were not likely to contribute appreciably to the flux of carbon between the two cell types. The amounts of ribulose-1,5-bisphosphate were high at low intercellular partial pressures of CO₂, and fell rapidly as the CO₂-assimilation rate increased with increasing intercellular partial pressures of CO₂, indicating that bundle-sheath CO₂ concentrations fell at low intercellular partial pressures of CO₂. In contrast, the amount of phosphoenolpyruvate and of C₄-cycle intermediates declined at low intercellular partial pressures of CO₂. This behaviour is discussed in relation to the co-ordination of carbon assimilation between the Calvin and C₄ cycles.Abbreviations PEP phosphoenolpyruvate - PGA glycerate-3-phosphate - p _i intercellular CO₂ pressure - RuBP ribulose-1,5-bisphosphate - triose-P triose phosphates     

Manifestation of the fragile site Xq27 in fibroblasts

Summary Fibroblasts from a heterozygous carrier for the Martin-Bell syndrome, who manifests the fragile site Xq27, were cloned to separate the population carrying the primary defect on the active X chromosome from the population with this defect on the inactive X. Clones with this defect on the active X manifest the fra(X)(q27) whereas clones from the other population are fra(X)-negative (Steinbach et al. 1983b). In this project, the replication status of the X chromosome manifesting the fra(X)(q27) was studied in seven clones with this defect on the active X.The results obtained on the clones were very similar to the results obtained from uncloned fibroblasts and lymphocytes. In the clones the fragile site was found manifested on the early replicating X in 73 cells and on the late replicating X in four cells.Since the defect is located on the active X chromosome of these cells the manifestation of the fragile site on the late replicating X suggests that the defect and the fragile site cannot be identical. It is concluded that there is no obligate synteny of this defect and the manifested fragile site.     

Opposing Effects of CREBBP Mutations Govern the Phenotype of Rubinstein-Taybi Syndrome and Adult SHH Medulloblastoma

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The metabolic capacity of lipid droplet localized acyl-CoA synthetase 3 is not sufficient to support local triglyceride synthesis independent of the endoplasmic reticulum in A431 cells

ACSL3 is the only long chain fatty acyl-CoA synthetase consistently found on growing and mature lipid droplets (LDs), suggesting that this specific localization has biological relevance. Current models for LD growth propose that triglycerides are synthesized by enzymes at the LD surface, with activated fatty acids provided by LD localized ACSL3, thus allowing growth independent of the ER. Here, we tested this hypothesis by quantifying ACSL3 on LDs from human A431 cells.RNAi of ACSL3 reduced the oleoyl-CoA synthetase activity by 83%, suggesting that ACSL3 is by far the dominant enzyme of A431 cells. Molar quantification revealed that there are 1.4 million ACSL3 molecules within a single cell. Metabolic labeling indicated that each ACSL3 molecule contributed a net gain of 3.1 oleoyl-CoA/s. 3D reconstruction of confocal images demonstrated that 530 individual lipid droplets were present in an average oleate fed A431 cell. A representative single lipid droplet with a diameter of 0.66?μm contained 680 ACSL3 molecules on the surface. Subcellular fractionation showed that at least 68% of ACSL3 remain at the ER even during extensive fatty acid supplementation. High resolution single molecule microscopy confirmed the abundance of cytoplasmic ACSL3 outside of LDs. Model calculations for triglyceride synthesis using only LD localized ACSL3 gave significant slower growth of LDs as observed experimentally.In conclusion, although ACSL3 is an abundant enzyme on A431 LDs, the metabolic capacity is not sufficient to account for LD growth solely by the local synthesis of triglycerides.     

Manufacturing development and clinical production of NKG2D chimeric antigen receptor–expressing T cells for autologous adoptive cell therapy

Background aims

Adoptive cell therapy employing natural killer group 2D (NKG2D) chimeric antigen receptor (CAR)-modified T cells has demonstrated preclinical efficacy in several model systems, including hematological and solid tumors. We present comprehensive data on manufacturing development and clinical production of autologous NKG2D CAR T cells for treatment of acute myeloid leukemia and multiple myeloma (ClinicalTrials.gov Identifier: NCT02203825). An NKG2D CAR was generated by fusing native full-length human NKG2D to the human CD3ζ cytoplasmic signaling domain. NKG2D naturally associates with native costimulatory molecule DAP10, effectively generating a second-generation CAR against multiple ligands upregulated during malignant transformation including MIC-A, MIC-B and the UL-16 binding proteins.

Assessing uncertainties in crop and pasture ensemble model simulations of productivity and N2O emissions

Simulation models are extensively used to predict agricultural productivity and greenhouse gas emissions. However, the uncertainties of (reduced) model ensemble simulations have not been assessed systematically for variables affecting food security and climate change mitigation, within multi‐species agricultural contexts. We report an international model comparison and benchmarking exercise, showing the potential of multi‐model ensembles to predict productivity and nitrous oxide (N₂O) emissions for wheat, maize, rice and temperate grasslands. Using a multi‐stage modelling protocol, from blind simulations (stage 1) to partial (stages 2–4) and full calibration (stage 5), 24 process‐based biogeochemical models were assessed individually or as an ensemble against long‐term experimental data from four temperate grassland and five arable crop rotation sites spanning four continents. Comparisons were performed by reference to the experimental uncertainties of observed yields and N₂O emissions. Results showed that across sites and crop/grassland types, 23%–40% of the uncalibrated individual models were within two standard deviations (SD) of observed yields, while 42 (rice) to 96% (grasslands) of the models were within 1 SD of observed N₂O emissions. At stage 1, ensembles formed by the three lowest prediction model errors predicted both yields and N₂O emissions within experimental uncertainties for 44% and 33% of the crop and grassland growth cycles, respectively. Partial model calibration (stages 2–4) markedly reduced prediction errors of the full model ensemble E‐median for crop grain yields (from 36% at stage 1 down to 4% on average) and grassland productivity (from 44% to 27%) and to a lesser and more variable extent for N₂O emissions. Yield‐scaled N₂O emissions (N₂O emissions divided by crop yields) were ranked accurately by three‐model ensembles across crop species and field sites. The potential of using process‐based model ensembles to predict jointly productivity and N₂O emissions at field scale is discussed.