Midbrain dopamine neuron differentiation: factors and fates

Dopaminergic neurons in the midbrain (mDNs) play a central role in the regulation of voluntary movement as well as other complex behaviors, and their loss is associated with Parkinson's disease (PD). The development of functional mDNs from multipotent progenitors is orchestrated by cell-intrinsic factors and cell-extrinsic environmental cues in a series of stages: early midbrain patterning, specification of mitotic precursors, postmitotic mDN development, and functional maturation. Of particular interest is how extracellular information is integrated with cell-intrinsic developmental programs. Cell fate mapping studies suggest that the stem-like progenitors for mDNs reside at the ventral midline floor plate, a region that also serves as a source of inductive signals for mDN specification such as Sonic Hedgehog (SHH). Cell replacement therapies, and in particular the use of embryonic or adult stem cell-derived dopaminergic neurons, offer potential novel treatment venues for PD, but such strategies require a detailed understanding of mDN development.     

Characterization of polar membrane lipids of the extremely halophilic bacterium Salinibacter ruber and possible role of cardiolipin

The lipid composition of the extremely halophilic bacterium Salinibacter ruber (Bacteroidetes) was investigated by thin layer chromatography, gas chromatography, high performance liquid chromatography and electrospray ionization-mass spectrometry. Polar lipids represent about 80% of the total lipid extract. The main polar lipids are a sulfonic acid analogue of ceramide (or capnine analogue), phosphatidylcholine, phosphatidylserine, dimethylphosphatidylethanolamine, phosphatidylglycerol, cardiolipin or bisphosphatidylglycerol, and a glycolipid. The major acyl chains in the phospholipids are C16:1 Delta9cis and C18:1 Delta11cis, while the sulfonolipid contains an amide-bound iso C15:0 fatty acid. On changing the salinity of the culture medium, no significant differences were found in the lipid profile or the unsaturation of the lipid fatty acyl chains. The structure of the cardiolipin, which represents 20% of polar lipids, has been elucidated by gas chromatography and electrospray ionization mass spectrometry analysis.     

Thiol‐disulfide organization in alliin lyase (alliinase) from garlic (Allium sativum)

Alliinase, an enzyme found in garlic, catalyzes the synthesis of the well-known chemically and therapeutically active compound allicin (diallyl thiosulfinate). The enzyme is a homodimeric glycoprotein that belongs to the fold-type I family of pyridoxal-5′-phosphate-dependent enzymes. There are 10 cysteine residues per alliinase monomer, eight of which form four disulfide bridges and two are free thiols. Cys368 and Cys376 form a S—S bridge located near the C-terminal and plays an important role in maintaining both the rigidity of the catalytic domain and the substrate-cofactor relative orientation. We demonstrated here that the chemical modification of allinase with the colored —SH reagent N-(4-dimethylamino-3,5-dinitrophenyl) maleimide yielded chromophore-bearing peptides and showed that the Cys220 and Cys350 thiol groups are accesible in solution. Moreover, electron paramagnetic resonance kinetic measurements using disulfide containing a stable nitroxyl biradical showed that the accessibilities of the two —SH groups in Cys220 and Cys350 differ. Neither enzyme activity nor protein structure (measured by circular dichroism) were affected by the chemical modification of the free thiols, indicating that alliinase activity does not require free —SH groups. This allowed the oriented conjugation of alliinase, via the —SH groups, with low- or high-molecular-weight molecules as we showed here. Modification of the alliinase thiols with biotin and their subsequent binding to immobilized streptavidin enabled the efficient enzymatic production of allicin.     

Isolation and functional expression of the bop gene from Halobiforma lacisalsi

A novel bop gene was described from Halobiforma lacisalsi strain AJ5^T, an extremely halophilic archaeon isolated from Ayakekum Lake, China. Following six rounds of PCR amplification based on the conserved fragment of the bop gene, the complete sequence of the bop gene, including the 5′ and 3′ flanking regions of the conserved fragment, was obtained by the ligation-mediated PCR amplification (LPA) approach. The data presented provide us with further insight into the distribution of bop-like genes in the family Halobacteriaceae. This is the first example of a bop-like gene in halophiles found in the high-pH environment. Alignment and hydropathy analysis of the deduced amino acid sequence identified the conserved functional sites as well as some variations compared with other bacterio-opsins. Molecular phylogenetic analysis revealed the position of the bacterio-opsin of strain AJ5, which is closest to that of Haloterrigena sp. arg-4 with 85% identity. In the presence of all-trans retinal, recombinant Escherichia coli cells expressing the gene turned dark purple. The purple membrane from the recombinant E. coli showed maximal absorption at 540 nm.     

Synthesis of the food flavoring methyl benzoate by genetically engineered Saccharomyces cerevisiae

Current means of production for plant-derived aroma compounds include chemical synthesis and extraction from plant material. Both methods are environmentally detrimental and relatively expensive: plant material is only seasonally available and only a small subset of the plant biomass produces the desired aroma compounds, while organic synthesis inevitably involves waste byproducts with a negative ecological impact. Benzenoids are a class of plant metabolites that includes a number of aroma compounds. This paper explores, for the first time, the feasibility of producing benzenoids in yeast. We present a method for the production of the phenylpropanoid methyl benzoate in Saccharomyces cerevisiae using benzoic acid as a substrate, by heterologous expression of Antirrhinum majus benzoic acid methyl transferase. Production was pH dependent with a maximal yield of approximately 50 microg of methyl benzoate per liter of culture per hour, and with linear kinetics over at least 24 h. In addition, we have analyzed two alternative expression vectors for the production of benzoic acid methyl transferase in S. cerevisiae: a constitutive triosephosphate isomerase promoter-based system was compared with a copper-inducible CUP1 promoter system. We find major differences in the amounts of methylbenzoate produced by these respective systems. Potential applications are discussed.     

Diabetic ulcers: The effect of thermal biofeedback-mediated relaxation training on healing

The present study was a preliminary investigation into the effect of thermal biofeedback-mediated relaxation training on healing of diabetic ulcers. Subjects were three males who had nonhealing diabetic ulcers of the toe, ankle, and leg, respectively. Treatment consisted of thermal biofeedback training at the ulcer site, handwarming training (in two of the three subjects), and home relaxation practice. Two subjects showed significant healing. Thermal biofeedback-mediated relaxation training may be a useful adjunct to traditional treatments for diabetic foot ulcers.The authors wish to acknowledge the assistance of Drs. Naomi Alazraki, Kathryn Morton, Louis Larsen, and Mr. Barry Burkinshaw of the Salt Lake VA Medical Center, without whom this study could not have been performed. The first author would also like to thank Dr. Edward Taub for his invaluable advice and support.     

Survival of Escherichia coli and Vibrio harveyi in Dead Sea water

Abstract When suspensions of Escherichia coli or the marine luminescent bacterium Vibrio harveyi were mixed with Dead Sea water, the number of viable bacteria decreased by 90% in a time varying from less than 1 h to several hours, depending on the bacterial strain tested. Survival was better at low temperatures, and diluting the Dead Sea water permitted prolonged survival of both coliform bacteria and V. harveyi . The death rate of E. coli in Dead Sea water was comparable to that in water from the Great Salt Lake (Utah). The high concentrations of calcium and magnesium in Dead Sea water, rather than the high total salinity, was identified as the main factor responsible for the rapid die-off. Exposure to direct solar irradiation significantly increased the die-off rate of E. coli in Dead Sea water.
Large numbers of coliform bacteria were recovered from the lake at distances of at least 20 m from a sewage discharge site on the western shore of the Dead Sea.     

Morphology and sedimentology of Halimeda bioherms from the eastern Java Sea (Indonesia)

Halimeda bioherms, occurring primarily along the western and southern margins of Kalukalukuang Bank in the eastern Java Sea, display a wide variety of thicknesses and shapes. In general, high-frequency forms of the northern bank are replaced by thicker and lower frequency forms along the deeper southern margin. Sidescan sonar data suggest aperiodic reworking of shallow bioherm crests of the northern bank into features suggestive of bedforms. These features are not associated with deeper bioherms of the southern bank. Cores from the bioherms indicate that they consist mostly of disarticulated Halimeda plates set in a lime mud matrix composed largely of Halimeda fragments and foraminifera tests. Carbon-14 dating shows that, with exception of some deep southern bank examples, bioherms are actively accreting. Results of mineralogy and elemental chemistry on piston core PC-12 suggest cyclic variations in Mg-calcite (cement in Halimeda utricles), which may be related to periodic excursions of cold Pacific throughflow water onto the bank. Composition of the Mg-calcite (8.6 mole-%) suggests a temperature of formation of about 22 °C, which is 7 °C below average surface water temperatures. The carbon and oxygen isotope compositions of both aragonite and Mg-calcite phases are remarkably homogeneous, but were inconclusive with regard to the cold-water intrusion hypothesis. However, a lack of reef-building corals below a depth of 15 m, abundance of Halimeda bioherms on the western margin of K-Bank, where upwelling is predicted, extensive boring of sedimentary particles by endolithic boring algae, and high nutrient values of water at the thermocline (50–70 m deep) all support the incursion of cold, nutrient-rich water onto the bank. Upwelling and nutrient overloading are suggested as explanations for remarkable algal growth at the expense of reef-building corals.     

An inventory of 13C abundances in coastal wetlands of Louisiana,USA: vegetation and sediments

Summary Organic carbon-rich sediments from the surface of fresh, intermediate, brackish and salt marshes of coastal Louisiana were sampled and analyzed for their ¹³C content. The average ¹³C from all sites within each wetland type was-27.8,-22.1,-16.9, and-16.2, for fresh, intermediate, brackish and salt marshes, respectively. Means from the fresh, intermediate and brackish marshes were significantly different at the 0.01 level. A mixing model using measurements of standing crop and ¹³C of plant carbon was applied to estimate the contribution of each species to the sedimentary carbon at four of the marsh sites. Sedimentary ¹³C values generally reflected that of the dominant species present at each site. Brackish and salt marsh samples, however, showed a negative shift of ¹³C with respect to whole plant carbon. We interpret these depeleted ¹³C values to be the result of more extensive organic matter decomposition and selective preservation of ¹³C-depleted refractory components in sediments from saline sites. The results of this study suggest that ¹³C composition of sedimentary carbon may offer a valuable tool for distinguishing subtle changes in paleohydrology of wetlands resulting from relative sea level changes.