Use of Radiation Hybrid panels to map genetic loci

The mapping of genetic loci within organisms has been accelerated by the advent of Radiation Hybrid (RH) panels. These panels are available for humans and non-humans including mice, baboon, rat, and canine. This article contains a general protocol for the use of the Genebridge 4 whole genome RH panel to map a human locus. This protocol may also be adjusted to suit the other RH panels currently available.     

Human impact on vegetation at the Alpine tree-line ecotone during the last millennium: lessons from high temporal and palynological resolution

Three mires and a small lake in the Swiss and Austrian Alps were studied palynologically at high resolution, covering the last 1,000, 400, 50 and 1,200 years, respectively. Methodological lessons include: (1) Sub-decadal resolution in upper, little-decomposed peat layers reveals recurrent marked fluctuations in both percentages and influx of regional tree-pollen types, reflecting variations in pollen production rather than in plant-population sizes. (2) Intermittent, single-spectrum pollen maxima in samples of sub-decadal resolution indicate pollen transport in clumps. This type of pollen transport may remain unrecognized in sections with lower sampling resolution, which may then lead to inappropriate interpretation in terms of plant-population sizes. (3) The detection of short-lived phases of human impact in decomposed peat requires sampling intervals as close as 0.2 cm. (4) PAR (pollen influx) may reflect vegetation dynamics more faithfully than percentages. Reliable PAR, however, is difficult to achieve in Alpine mires due to past human impact on peat growth, even when complex depth–age modelling techniques are used. Critical comparison of PAR with percentages is therefore essential. (5) Careful consideration of spatial scales in pollen signals (local–regional and subdivisions) is essential for a realistic palaeo-ecological interpretation. Results in terms of past human impact on vegetation are summarized as follows: (1) Trends in pollen types reflecting regional human action are in general agreement with earlier findings for the western Swiss Alps, allowing for regional differences. (2) All mires in the Alps investigated here and in an earlier study experienced human impact during the last millennium. The studied small lake, lying in sub-alpine pasture, records forest dynamics at a lower elevation since a.d. 800.     

In vitro inhibition of vitamin K dependent carboxylation by tetrachloropyridinol and the imidazopyridines

The compounds 2,3,5,6-tetrachloro-4-pyridinol (TCP) and the structurally related imidazopyridines (IP) cause hemorrhage and lower the plasma prothrombin level in animals. In vitro, TCP and the IP are more potent inhibitors of both the vitamin K dependent carboxylase which catalyzes the posttranslational gamma-carboxylation of specific glutamyl residues in proteins and the related vitamin K epoxidase activity than they are either of vitamin K epoxide reductase or of NAD-(P)H-K oxidoreductase. TCP and IP, as is the case with the coumarin and indandione anticoagulants, are competitive inhibitors of NAD(P)H-K oxidoreductae with respect to NADH. The epoxide reductase from coumarin-resistant rats is quite resistant to inhibition not only by warfarin but also by the IP, and to a lesser extent by TCP. When interpreted in light of published in vivo experiments, the data suggest that the principal site of anticoagulant action of the IP, but not TCP, is the epoxide reductase. The anticoagulant effect of TCP may be inhibition of the carboxylase itself. TCP is a significantly more potent inhibitor of the carboxylase and epoxidase than the IP; it inhibits both the enzymatic activities to the same degree with 50% inhibition observed at about 10(-5) M. Inhibition of the carboxylase by TCP is not competitive with respect to the pentapeptide substrate phenylalanyl-leucylglutamylglutamylleucine nor with respect to the following components of the in vitro carboxylase assay: imidazole, pyridoxal 5'-phosphate, dithiothreitol, KCl, sodium bicarbonate, oxygen, and vitamin K. The order of addition of components of the assay relative to the addition of inhibitor did not affect the degree of inhibition. Inhibition is readily reversed in experiments designed to dissociate an enzyme-inhibitor complex. Analysis of double-inhibitor experiments suggests that TCP and IP have the same binding site on the carboxylase.     

Effect of solvent viscosity on the heme-pocket dynamics of photolyzed (carbonmonoxy)hemoglobin

The heme-pocket dynamics subsequent to carbon monoxide photolysis from human hemoglobin have been monitored as a function of glycerol-water solvent composition with time-resolved resonance Raman spectroscopy. Prompt (geminate) ligand recombination rates and the transient heme-pocket geometry established within 10 ns after photolysis appear to be largely independent of solvent composition. The rate of relaxation of the transient geometry to an equilibrium deoxy configuration is, however, quite sensitive to solvent composition. These observations suggest that the former processes result from local, internal motions of the protein, while the relaxation dynamics of the proximal heme pocket are predicated upon more global protein motions that are dependent upon solvent viscosity.     

GROWTH AND DEVELOPMENT OF THE MALE GAMETOPHYTE OF GINKGO BILOBA WITHIN THE OVULE (IN VIVO)

The mature male gametophyte of Ginkgo biloba can be divided into two regions: a large saccate structure that is suspended within the fertilization chamber above the archegonia, and a pervasive, highly branched haustorial system that ramifies through the intercellular air spaces of the apex of the nucellus. This morphology appears to differ in many ways from the simpler more typical male gametophytes of most other groups of seed plants. Growth and development of the male gametophyte of Ginkgo biloba were studied using computer reconstruction techniques to generate images of the gametophyte from data derived from serial sections through the ovule. These investigations reveal that morphological development of the male gametophyte of Ginkgo biloba is divided into three distinct phases: 1) Germination, characterized by an initial brief period of diffuse growth. This phenomenon has not been described for any other seed plant male gametophyte; 2) Initiation of tip growth and the formation of a tubular body, as typifies all seed plant male gametophytes. In Ginkgo, this is accompanied by a high degree of branching, giving rise to an extensively branched haustorial system; 3) Late swelling of the proximal unbranched portion of the gametophyte resulting in formation of the saccate structure that is characteristic of the mature gametophyte. This process appears to be very similar to late development in cycad male gametophytes. Thus, despite the seemingly anomalous morphology of the mature male gametophyte of Ginkgo biloba, specific patterns of growth and development are in many ways similar to growth processes expressed by the male gametophytes of some or all major groups of seed plants.     

Selected contribution: effects of ischemia-reperfusion on vascular contractility and alpha(1)-adrenergic-receptor signaling in the rat tail artery.

To determine the effects of ischemia-reperfusion (I/R) on alpha(1)-adrenergic-receptor (alpha(1)-AR) functions, alpha(1)-AR-mediated contraction, inositol phosphate (IP) accumulation, and alpha(1)-AR-G protein coupling were examined in the tail arteries of anesthetized rats after 60 min of ischemia and 60 min of reperfusion. The contractile response to norepinephrine (NE) was significantly increased after I/R, whereas the contractile response to KCl remained unchanged. This was accompanied by a 69% increase in NE-stimulated IP accumulation. Furthermore, receptor-stimulated coupling of alpha(1a)-AR to G alpha(q/11) proteins was increased, whereas the coupling of alpha(1b)-AR or alpha(1d)-AR to their G proteins was not altered by I/R. These changes in vascular alpha(1)-AR function occurred without concurrent alteration in expression levels of membrane alpha(1)-AR subtypes or in the associated G proteins. These data demonstrate that I/R increases alpha(1a)-AR-G(q/11) protein coupling and alpha(1)-AR-stimulated IP accumulation in the tail artery. The alterations in alpha(1)-AR signaling are associated with and may underlie the enhanced contractile response of the tail artery to adrenergic stimulation after I/R.