Enhanced levels of methionine and cysteine in transgenic alfalfa (Medicago sativa L.) plants over-expressing the Arabidopsis cystathionine gamma-synthase gene

With the aim of increasing the methionine level in alfalfa (Medicago sativa L.) and thus improving its nutritional quality, we produced transgenic alfalfa plants that expressed the Arabidopsis cystathionine gamma-synthase (AtCGS), the enzyme that controls the synthesis of the first intermediate metabolite in the methionine pathway. The AtCGS cDNA was driven by the Arabidopsis rubisco small subunit promoter to obtain expression in leaves. Thirty transgenic plants were examined for the transgene protein expression, and four lines with a high expression level were selected for further work. In these lines, the contents of methionine, S-methylmethionine (SMM), and methionine incorporated into the water-soluble protein fraction increased up to 32-fold, 19-fold, and 2.2-fold, respectively, compared with that in wild-type plants. Notably, in these four transgenic lines, the levels of free cysteine (the sulphur donor for methionine synthesis), glutathione (the cysteine storage and transport form), and protein-bound cysteine increased up to 2.6-fold, 5.5-fold, and 2.3-fold, respectively, relative to that in wild-type plants. As the transgenic alfalfa plants over-expressing AtCGS had significantly higher levels of both soluble and protein-bound methionine and cysteine, they may represent a model and target system for improving the nutritional quality of forage crops.     

Herbicide-resistance conferred by expression of a catalytic antibody in Arabidopsis thaliana

Engineering herbicide resistance in crops facilitates control of weed species, particularly those that are closely related to the crop, and may be useful in selecting lines that have undergone multiple transformation events. Here we show that herbicide-resistant plants can be engineered by designing an herbicide and expressing a catalytic antibody that destroys the herbicide in planta. First, we developed a carbamate herbicide that can be catalytically destroyed by the aldolase antibody 38C2. This compound has herbicidal activity on all three plant species tested. Second, the light chain and half of the heavy chain (Fab) of the catalytic antibody were targeted to the endoplasmic reticulum in two classes of Arabidopsis thaliana transformants. Third, the two transgenic plants were crossed to produce an herbicide-resistant F1 hybrid. The in vitro catalytic activity of the protein from F1 hybrids corroborates that catalytic antibodies can be constitutively expressed in transgenic plants, and that they can confer a unique trait.     

Energy-based stochastic control of neural mass models suggests time-varying effective connectivity in the resting state

Several studies posit energy as a constraint on the coding and processing of information in the brain due to the high cost of resting and evoked cortical activity. This suggestion has been addressed theoretically with models of a single neuron and two coupled neurons. Neural mass models (NMMs) address mean-field based modeling of the activity and interactions between populations of neurons rather than a few neurons. NMMs have been widely employed for studying the generation of EEG rhythms, and more recently as frameworks for integrated models of neurophysiology and functional MRI (fMRI) responses. To date, the consequences of energy constraints on the activity and interactions of ensembles of neurons have not been addressed. Here we aim to study the impact of constraining energy consumption during the resting-state on NMM parameters. To this end, we first linearized the model, then used stochastic control theory by introducing a quadratic cost function, which transforms the NMM into a stochastic linear quadratic regulator (LQR). Solving the LQR problem introduces a regime in which the NMM parameters, specifically the effective connectivities between neuronal populations, must vary with time. This is in contrast to current NMMs, which assume a constant parameter set for a given condition or task. We further simulated energy-constrained stochastic control of a specific NMM, the Wilson and Cowan model of two coupled neuronal populations, one of which is excitatory and the other inhibitory. These simulations demonstrate that with varying weights of the energy-cost function, the NMM parameters show different time-varying behavior. We conclude that constraining NMMs according to energy consumption may create more realistic models. We further propose to employ linear NMMs with time-varying parameters as an alternative to traditional nonlinear NMMs with constant parameters.     

Similarities between Exogenously- and Endogenously-Induced Envelope Stress: The Effects of a New Antibacterial Molecule,TPI1609-10

Antibiotics with novel and/or multiple targets are highly desirable in the face of the steady rise of clinical antibiotic resistance. We have screened and identified small molecules, typified by the compound TPI1609-10 (aka SM10), with antibiotic activity against both gram-positive and gram-negative bacteria. SM10 was screened in vitro to bind branched Holliday junction intermediates of homologous recombination and tyrosine recombinase-mediated recombination; thus, the cellular targets of the small molecules were expected to include the RuvABC Holliday junction resolvasome and the XerCD complex involved in proper segregation of replicated chromosomes to daughter cells. SM10 indeed induces DNA damage and filamentation in E. coli. However, SM10 also induces envelope stress and causes increased production of intracellular reactive oxygen species. In addition, SM10 has similar effects to endogenously-induced envelope stress via overproducing outer membrane proteins (OmpC and OmpF), which also induces the SOS response, chromosome fragmentation, and production of reactive oxygen species. The synergy between SM10, and cerulenin, a fatty acid synthesis inhibitor, together with the SM10 hypersensitivity of cpx and rpoE mutants, further support that SM10''s mode of action damages membrane damage. The lethality of SM10 treatment and of OmpC overproduction are observed in both aerobically- and anaerobically-grown cells, and is accompanied by substantial DNA damage even anaerobically. Thus, only some DNA damage is due to reactive oxygen. We propose that membrane depolarization and the potential reduction in intracellular pH, leading to abasic site formation, cause a substantial amount of the DNA damage associated with both SM10 treatment and endogenous envelope stress. While it is difficult to completely exclude effects related to envelope damage as the sources of DNA damage, trapping intermediates associated with DNA repair and chromosome segregation pathways remains very likely. Thus SM10 may have distinct but synergistic modes of action.     

Novel terpenoids of the fungus Aspergillus insuetus isolated from the Mediterranean sponge Psammocinia sp. collected along the coast of Israel

Three novel meroterpenoids, insuetolides A-C (1-3) and four drimane sesquiterpenes, the new (E)-6-(4'-hydroxy-2'-butenoyl)-strobilactone A (4) and the known 2α, 9α, 11-trihydroxy-6-oxodrim-7-ene (5), strobilactone A (6) and (E,E)-6-(6',7'-dihydroxy-2',4'-octadienoyl)-strobilactone A (7), were isolated from the EtOAc extract of the culture medium of the marine-derived fungus Aspergillus insuetus (OY-207), which was isolated from the Mediterranean sponge Psammocinia sp. The structures of the compounds were determined by spectroscopic methods. Insuetolides A-C reveal a new carbon skeleton derived from the cyclization of farnesyl and 3, 5-dimethylorsellinic acid. Compounds 1, 6, and 7 exhibited anti-fungal activity towards Neurospora crassa with MIC values of 140, 242, and 162 μM, respectively; and compounds 3, 4, and 7 exhibited mild cytotoxicity towards MOLT-4 human leukemia cells.     

Movement of protein and macromolecules between host plants and the parasitic weed Phelipanche aegyptiaca Pers.

Little is known about the translocation of proteins and other macromolecules from a host plant to the parasitic weed Phelipanche spp. Long-distance movement of proteins between host and parasite was explored using transgenic tomato plants expressing green fluorescent protein (GFP) in their companion cells. We further used fluorescent probes of differing molecular weights to trace vascular continuity between the host plant and the parasite. Accumulation of GFP was observed in the central vascular bundle of leaves and in the root phloem of transgenic tomato plants expressing GFP under the regulation of AtSUC2 promoter. When transgenic tomato plants expressing GFP were parasitized with P. aegyptiaca, extensive GFP was translocated from the host phloem to the parasite phloem and accumulated in both Phelipanche tubercles and shoots. No movement of GFP to the parasite was observed when tobacco plants expressing GFP targeted to the ER were parasitized with P. aegyptiaca. Experiments using fluorescent probes of differing molecular weights to trace vascular continuity between the host plant and the parasite demonstrated that Phelipanche absorbs dextrans up to 70 kDa in size from the host and that this movement can be bi-directional. In the present study, we prove for the first time delivery of proteins from host to the parasitic weed P. aegyptiaca via phloem connections, providing information for developing parasite resistance strategies.     

Cholesterol uptake is dependent on membrane fluidity in mycoplasmas

The transfer of elaidate-enriched Acholeplasma laidlawii cells in culture from 37°C to 4°C virtually arrested exogenous cholesterol incorporation into the cell membrane. Cholesterol uptake continued, though at a slower rate, in oleate-enriched A. laidlawii cells undergoing similar temperature shift-down. It is concluded that the incorporation of exogenous cholesterol into the cell membrane of living mycoplasmas is rapid when the membrane lipid bilayer is in the liquid-crystalline state and very slow when the lipid bilayer is in the gel state.