Cells that express MyoD mRNA in the epiblast are stably committed to the skeletal muscle lineage

The epiblast of the chick embryo contains cells that express MyoD mRNA but not MyoD protein. We investigated whether MyoD-positive (MyoDpos) epiblast cells are stably committed to the skeletal muscle lineage or whether their fate can be altered in different environments. A small number of MyoDpos epiblast cells were tracked into the heart and nervous system. In these locations, they expressed MyoD mRNA and some synthesized MyoD protein. No MyoDpos epiblast cells differentiated into cardiac muscle or neurons. Similar results were obtained when MyoDpos cells were isolated from the epiblast and microinjected into the precardiac mesoderm or neural plate. In contrast, epiblast cells lacking MyoD differentiated according to their environment. These results demonstrate that the epiblast contains both multipotent cells and a subpopulation of cells that are stably committed to the skeletal muscle lineage before the onset of gastrulation. Stable programming in the epiblast may ensure that MyoDpos cells express similar signaling molecules in a variety of environments.     

Analysis of neurosterols and neurosteroids by mass spectrometry

In man the brain represents about 2% of the body weight, but contains 25% of the body's cholesterol. Cholesterol itself does not cross the blood-brain barrier and is synthesised in situ. Excess cholesterol from brain is exported in the form of oxysterols, or metabolised to steroids, which in contrast to cholesterol can cross the blood-brain barrier. Steroids and oxysterols may be synthesised in brain, but can also be transported into brain from peripheral tissue. Both oxysterols and steroids have biological activity in brain. They can behave as ligands for classical nuclear receptors, and exert their effects over hours to days, or interact with neurotransmitter gated ion channels and modulate neural transmission exerting their effects in milliseconds. The exact sterol and steroid content of brain has yet to be thoroughly characterised. In this mini-review we will discuss mass spectrometry methods for the analysis of steroids and sterols in brain, and propose methods suitable for the profiling of different brain regions with high sensitivity (sub pg) and specificity.     

Construction of polyamine-modified uridine and adenosine derivatives--evaluation of DNA binding capacity and cytotoxicity in vitro

We here report the synthesis of the two polyamine-based nucleoside derivatives 5-{[bis-(3-aminopropyl)amino]acetamido-1-propynyl}uridine and 2-{[bis-(3-aminopropyl)amino]-acetamido-1-propynyl}adenosine. The various polyamine derivatives have been used in thermal melting analysis using DNA from herring testes, and in cellular studies using four different cell lines. The compounds were all found to be non-toxic, thus holding good promise for future use as siRNA building blocks.     

Dark diversity in dry calcareous grasslands is determined by dispersal ability and stress‐tolerance

Temperate calcareous grasslands are characterized by high levels of species richness at small spatial scales. Nevertheless, many species from a habitat‐specific regional species pool may be absent from local communities and represent the ‘dark diversity’ of these sites. Here we investigate dry calcareous grasslands in northern Europe to determine what proportion of the habitat‐specific species pool is realized at small scales (i.e. how the community completeness varies) and which mechanisms may be contributing to the relative sizes of the observed and dark diversity. We test whether the absence of particular species in potentially suitable grassland sites is a consequence of dispersal limitation and/or a low ability to tolerate stress (e.g. drought and grazing). We analysed a total of 1223 vegetation plots (1 × 1 m) from dry calcareous grasslands in Sweden, Estonia and western Russia. The species co‐occurrence approach was used to estimate the dark diversity for each plot. We calculated the maximum dispersal distance for each of the 291 species in our dataset by using simple plant traits (dispersal syndrome, growth form and seed characteristics). Large seed size was used as proxy for small seed number; tall plant height and low S‐strategy type scores were used to characterise low stress‐tolerance. Levels of small‐scale community completeness were relatively low (more species were absent than present) and varied between the grasslands in different geographic areas. Species in the dark diversity were generally characterized by shorter dispersal distances and greater seed weight (fewer seeds) than species in the observed diversity. Species within the dark diversity were generally taller and had a lower tolerance of stressful conditions. We conclude that, even if temperate grasslands have high levels of small‐scale plant diversity, the majority of potentially suitable species in the regional species pool may be absent as a result of dispersal limitation and low stress‐tolerance.     

The Enzyme and the cDNA Sequence of a Thermolabile and Double-Strand Specific DNase from Northern Shrimps (Pandalus borealis)

Background

We have previously isolated a thermolabile nuclease specific for double-stranded DNA from industrial processing water of Northern shrimps (Pandalus borealis) and developed an application of the enzyme in removal of contaminating DNA in PCR-related technologies.

Methodology/Principal Findings

A 43 kDa nuclease with a high specific activity of hydrolysing linear as well as circular forms of DNA was purified from hepatopancreas of Northern shrimp (Pandalus borealis). The enzyme displayed a substrate preference that was shifted from exclusively double-stranded DNA in the presence of magnesium to also encompass significant activity against single-stranded DNA when calcium was added. No activity against RNA was detected. Although originating from a cold-environment animal, the shrimp DNase has only minor low-temperature activity. Still, the enzyme was irreversibly inactivated by moderate heating with a half-life of 1 min at 65°C. The purified protein was partly sequenced and derived oligonucleotides were used to prime amplification of the encoding cDNA. This cDNA sequence revealed an open reading frame encoding a 404 amino acid protein containing a signal peptide. By sequence similarity the enzyme is predicted to belong to a family of DNA/RNA non-specific nucleases even though this shrimp DNase lacks RNase activity and is highly double-strand specific in some respects. These features are in agreement with those previously established for endonucleases classified as similar to the Kamchatka crab duplex-specific nuclease (Par_DSN). Sequence comparisons and phylogenetic analyses confirmed that the Northern shrimp nuclease resembles the Par_DSN-like nucleases and displays a more distant relationship to the Serratia family of nucleases.

Conclusions/Significance

The shrimp nuclease contains enzyme activity that may be controlled by temperature or buffer compositions. The double-stranded DNA specificity, as well as the thermolabile feature, strengthens its potential for in vitro applications.     

The effect of irradiation by a He-Ne laser and phytohemagglutinin on lymphocyte mitochondria

Electron-microscopic morphometry has been applied to study mitochondria on ultrathin sections of lymphocytes from human peripheral blood. It has been shown that the stimulation of lymphocytes by the mitogen phytohemagglutinin (PHA) 1 h causes increases in the quantity of mitochondria per cellular section (17%) as well as in the total area of mitochondria per cell section (35%), i.e. an increase in mitochondrial mass. Taking into account known facts about growth and division of mitochondria in late phases of cellular cycle, one can suppose that described above changes in mitochondria during G0----G1 transition under action of PHA belong to an early phase of biogenesis of mitochondria. In the contrary, irradiation of lymphocytes with He-Ne-laser (lambda = 632.8 nm) in dose 56 J/m2 which does not cause the G0----C1 transition, results in the increase in the number of mitochondria per cellular section (20%) but not increase in the total area of mitochondria per cell section. The last finding indicates to some modification of space configuration of the mitochondria without any changes in their mass. The increase in the quantity of mitochondria per cellular section after the irradiation could be related with the increase in electrochemical proton gradient and in phosphorylating activity of mitochondria. He-Ne-laser radiation as well as mitogen PHA cause some deaggregation of mitochondria (this is more pronounced in case of PHA) which may be related to their functional activation.     

Action of the radiation from a neon laser and from noncoherent blue light on Escherichia coli bacteria

It was shown that under defined conditions blue light can accelerate E. coli WP2 growth. The stimulatory effect is a function of radiation dose, intensity wave length, and postirradiation incubation time.