A simple method of characterizing mitochondrial ribonucleic acid.

A simple method of isolating and characterizing RNA from L-cell mitochondria is described. The mitochondrial fraction is lysed by sodium dodecyl sulphate, and the RNA fractionated by sucrose-density-gradient centrifugation. The efficacy of proteinase K in preventing ribonuclease activity is also demonstrated.     

Testing and characterizing the two-stage model of carcinogenesis for a wide range of human cancers

The age dependence of incidence for 45 cancer types in three populations is analyzed on a two-stage kinetic model containing three determinable parameters: (i) the fraction of population at risk for a cancer; (ii) the product of the frequencies of cancer-producing mutations, and (iii) the growth rate of the transformed clone from which a cancer ultimately evolves. The model, simplifying that proposed by others, fits many cancers. Data are easily handled in terms of the derived parameters, providing the basis for epidemiological analysis, here applied in detail to liver, cervix and testis cancers for nearly 50 world-wide populations. We identify 12 cancers for which only a limited fraction of the population is at risk. We argue that the appearance of most cancers requires at least three mutational events. For child, youth, or early adult cancers, one mutation may be congenital. We arrange the various cancers in a descending scale, defining six groups that differ in the derived mutation frequencies. A cancer appearing later in life, for which the whole population can be at risk, shows a low mutation frequency, consistent with background spontaneous mutation. The other cancers require increases in mutation frequency, arising from an increased rate of cell division and/or mutation rate.     

Testing simple indices of habitat proximity

Simple measures of habitat proximity made primarily on the basis of land cover are widely used in the ecological literature to infer habitat connectivity, or the potential for animal movement among resource patches. However, such indices rarely have been tested against observations of animal movement or against more detailed biological models. We developed a priori expectations as to the types of study systems and organisms for which various habitat proximity indices would be best suited. We then used data from three study systems and four species to test which, if any, of the indices were good predictors of population-level responses. Our a priori expectations about index performance were not upheld. The indices that consider both habitat area and distance from the focal patch were highly correlated with each other, suggesting that they do index similar quantities. However, none of the indices performed well in predicting population response variables. The results suggest that the pattern of habitat cover alone may be insufficient to predict the process of animal movement.     

How the kinetic parameters of the simple carrier are affected by an applied voltage

The kinetic parameters ($\text{K, R}{}_{\mathrm{<mtext>oo</mtext>}}\text{, R}{}_{12}\text{, R}{}_{21}\text{,}\text{and}\text{R}{}_{\mathrm{<mtext>ee</mtext>}}$) of the simple carrier model are analysed as a function of applied voltage for various cases of charged substrates and carriers. If certain parameters are invariant with voltage, strong conclusions can be made as to the charge on the free carrier. In one particular case (where the parameter $\text{R}{}_{00}$ is invariant while $\text{R}{}_{12}$ and $\text{R}{}_{21}$ display a nonlinear dependence on the exponential of the voltage) it may be possible to identify the isomerisation of the carrier-substrate complex.     

Testing a simple stochastic model for the dynamics of waterfowl aggregations

We consider a simple stochastic model for the dynamics of mixed-species waterfowl aggregations and describe two methods for assessing the fit of this model to field data. The model does not incorporate species-specific behavior. It assumes that all birds act independently and incorrectly predicts an exponential distribution for inter-event times. We reject this model, show that 29% of the birds move in groups of two or more birds, and demonstrate that the distribution of inter-event times between the movements of groups of birds is exponential. We find no difference in movement rates or group sizes between seasons, and no difference between groups arriving into or departing from the observed aggregations. An analysis of group composition suggests that species at low abundance behave differently than species at high abundance: birds with few conspecifics are more likely to move in mixed-species groups than birds with many conspecifics. We suggest that simple stochastic models provide a useful way to explore the dynamics of animal behavior.     

Testing a simple rule for dominance in resource competition

Competition for limiting resources long has been considered an important factor generating community structure. A minimal model of resource competition predicts that the species that reduces the limiting resource R to the lowest level ([Formula: see text]) will exclude its competitors. Whether this "[Formula: see text] rule" is robust to violations of model assumptions remains largely unknown. I conducted a competition experiment with four species of bacterivorous protists in laboratory microcosms and predicted the outcome from each species' [Formula: see text] value. I also examined how the outcome of competition, species abundances, and the effect of protists on bacterial density varied with productivity. Microcosms were unstirred batch cultures containing a variety of bacteria, challenging the robustness of the simplest competition models. Protists with low [Formula: see text] values were less affected by competition, although competing protists often coexisted. The values of [Formula: see text] can predict competitive dominance, even in the absence of competitive exclusion. Other model predictions were less robust. Contrary to expectation, densities of grazed bacteria increased with productivity, and the effect of some protists on bacterial density did not vary with productivity. Bacterial heterogeneity may account for deviations from model predictions. Further experiments should examine the conditions under which simple rules can be expected to identify dominant species.     

Identifying and characterizing the joint cavity-forming cell

For many years, a large body of circumstantial evidence supported the notion that the synovial membrane produced the hyaluronan-rich synovial fluid. A quantitative cytochemical technique for uridine-diphospho glucose dehydrogenase (UDPGD) activity established that fibroblast-like cells on the intimal surface of the synovial lining made a specific contribution to maintaining these glycosaminoglycan levels. Our studies have aimed to determine the mechanisms that control the attainment and persistence of this differentiated phenotype, and have recently focused on their appearance during joint cavity development in the embryonic limb; a process that is dependent upon skeletal movement. These in situ micro-biochemical studies have shown that cells bordering the presumptive joint cavity exhibit raised UDPGD activity, are associated with a matrix rich in hyaluronan and show immobilization-induced loss in such characteristics. Together with complimentary studies in adult joints, this suggests that mechanical stimuli promote the acquisition of this joint line-forming phenotype. For this reason our studies have attempted to identify the 'up-stream' mechano-dependent factors that control these events. Endothelial cells respond to mechanical stimuli by activating, via phosphorylation, mitogen activated protein kinase/extracellular signal-regulated kinase (MAPkinase/ERK). Using phospho-specific anti-ERK-1/2 antibodies we have shown that immunolabelling of developing limbs shows a clear joint line-selective activation during cavitation, with little if any labelling within neighbouring elements, and that this is abolished in immobilized limbs. In an attempt to facilitate the final mechanistic deciphering of these responses we have used an in vitro-based approach and found by Western blotting that active ERK-1/2 expression was increased in cultured articular surface cells following application of dynamic mechanical strain. Intriguingly, the use of a selective inhibitor (PD98059) of ERK activation by its classical activating kinase, Mek, to restrict such strain-induced increases, produced an enhanced strain-related increase in UDPGD mRNA expression. This suggests that mechano-dependent ERK activation serves a feedback regulatory role during differentiation of these cells. Whilst it is clear that these in vitro experiments serve a useful function, it is clear that they generally take little regard of the influence that might be provided by cell-cell and cell-matrix interactions within the developing limb's complex and dynamic environment and architecture. It is therefore imperative that we attempt to bridge the gap between the cell biology of such phenomena on the one hand, and the morphological approach to this same problem on the other.     

A simple experimental approach to the Determination of carrier transport parameters for unlabeled substrate analogs

A method is described by which affinities and transport rates for unlabeled substrate analogs are readily determined, and which is based on the effect of an unlabeled analog upon the rate of transport of a labeled substrate present at a low concentration on the trans side of the membrane. The procedure is widely applicable since it does not depend on assumptions about rate-limiting steps and holds for both active and non-active systems. Here it is applied in an experimental study of the facilitated diffusion system for choline in erythrocytes, and it is shown that the transport parameters for a test substrate obtained by this method are the same as those found when the transport of the substrate is followed directly.     

Functional expression of the CMP-sialic acid transporter in Escherichia coli and its identification as a simple mobile carrier

The architectural conservation of nucleotide sugar transport proteins (NSTs) enabled the theoretical prediction of putative NSTs in diverse gene databases. In the human genome, 17 NST sequences have been identified but only six have been unequivocally characterized with respect to their transport specificities. Defining transport characteristics of recombinant NSTs has become a major challenge because true zero background systems are widely absent. Production of recombinant NSTs in heterologous systems has developed multifunctionality for some NSTs leading to a novel level of complexity in the field. Assuming that (1) the specificity of NSTs is determined at the primary sequence level and (2) the proteins are autonomously functional units, final definition of the substrate specificity will depend on the use of isolated transport proteins. Herein, we describe the first report of the functional expression of mouse CMP-sialic acid transporter (CST) in Escherichia coli and thus provide significant progress towards the production of transporter proteins in quantities suitable for functional and structural analyses. Recovery of the active NST from inclusion bodies was achieved after solubilization with 8 M urea and stepwise renaturation. After reconstitution into phospholipid vesicles, the recombinant protein demonstrated specific transport for CMP-N-acetylneuraminic acid (CMP-Neu5Ac) with no transport of UDP-sugars. Kinetic studies carried out with CMP-Neu5Ac and established CMP-Neu5Ac antagonist's evaluated natural conformation of the reconstituted protein and clearly demonstrate that the transporter acts as a simple mobile carrier.     

A simple experimental approach to the determination of carrier transport parameters for unlabeled substrate analogs.

A method is described by which affinities and transport rates for unlabeled substrate analogs are readily determined, and which is based on the effect of an unlabeled analog upon the rate of transport of a labeled substrate present at a low concentration on the trans side of the membrane. The procedure is widely applicable since it does not depend on assumptions about rrate-limiting steps and holds for both active and non-active systems. Here it is applied in an experimental study of the facilitated diffusion system for choline in erythrocytes, and it is shown that the transport parameters for a test substrate obtained by this method are the same as those found when the transport of the substrate is followed directly.     

Defining and characterizing high-rainfall Mediterranean climates

Abstract

The world's five Mediterranean-climate regions are typified by similarities in climate reflected in structural vegetation patterns. We developed a methodology to define high-rainfall (annual precipitation ≥800 mm) Mediterranean climates and contrast patterns of between- and within-region climatic heterogeneity. Climatic indices describing the seasonality of the climate were computed in Geographic Information Systems and included in a numerical analysis, in addition to selected bioclimatic variables, using the pattern analysis program PATN. Selected locations from high-rainfall Mediterranean-climate regions were grouped according to climatic similarity, and within-region climatic heterogeneity was compared between regions. Areas traditionally not considered as Mediterranean-climate regions emerged in the classification. Only the Mediterranean-type systems within the Pacific north-west of North America and south-central Chile comprised a substantial extent of high-rainfall Mediterranean climate. Both of these regions were characterized by a high climatic diversity and steep topographic gradients. The numerical analysis demonstrated the high diversity of the high-rainfall Mediterranean-climate type and differences in within-region climatic heterogeneity between the world's high-rainfall Mediterranean-climate regions.