Elemental distribution in striated muscle and the effects of hypertonicity: Electron probe analysis of cryo sections

A method of rapid freezing in supercooled Freon 22 (monochlorodifluoromethane) followed by cryoultramicrotomy is described and shown to yield ultrathin sections in which both the cellular ultrastructure and the distribution of diffusible ions across the cell membrane are preserved and intracellular compartmentalization of diffusabler ions can be quantitated. Quantitative electron probe analysis (Shuman, H., A.V. Somlyo, and A.P. Somlyo. 1976. Ultramicros. 1:317-339.) of freeze-dried ultrathin cryto sections was found to provide a valid measure of the composition of cells and cellular organelles and was used to determine the ionic composition of the in situ terminal cisternae of the sarcoplasmic reticulum (SR), the distribution of CI in skeletal muscle, and the effects of hypertonic solutions on the subcellular composition if striated muscle. There was no evidence of sequestered CI in the terminal cisternae of resting muscles, although calcium (66mmol/kg dry wt +/- 4.6 SE) was detected. The values of [C1](i) determined with small (50-100 nm) diameter probes over cytoplasm excluding organelles over nuclei or terminal cisternae were not significantly different. Mitochondria partially excluded C1, with a cytoplasmic/ mitochondrial Ci ratio of 2.4 +/- 0.88 SD. The elemental concentrations (mmol/kg dry wt +/- SD) of muscle fibers measured with 0.5-9-μm diameter electron probes in normal frog striated muscle were: P, 302 +/- 4.3; S, 189 +/- 2.9;C1, 24 +/- 1.1;K, 404 +/- 4.3, and Mg, 39 +/- 2.1. It is concluded that: (a) in normal muscle the "excess CI" measured with previous bulk chemical analyses and flux studies is not compartmentalized in the SR or in other cellular organelles, and (b) the cytoplasmic C1 in low [K](0) solutions exceeds that predicted by a passive electrochemical distribution. Hypertonic 2.2 X NaCl, 2.5 X sucrose, or 2.2 X Na isethionate produced: (a) swollen vacuoles, frequently paired, adjacent to the Z lines and containing significantly higher than cytoplasmic concentrations of Na and Cl or S (isethionate), but no detectable Ca, and (b) granules of Ca, Mg, and P = approximately (6 Ca + 1 Mg)/6P in the longitudinal SR. It is concluded that hypertonicity produces compartmentalized domains of extracellular solutes within the muscle fibers and translocates Ca into the longitudinal tubules.     

Multiple mRNAs are generated from the chicken lysozyme gene

We have determined the DNA sequence of a 770 by Pst I fragment containing 450 nucleotides of the 5′ flanking region of the chicken lysozyme gene. S1-nuclease mapping was performed to localize the 5′ end of nuclear RNA containing lysozyme-specific sequences and of the mRNA. We present evidence that the 5′ noncoding region of the chicken lysozyme mRNA is heterogeneous in length. The 5′ termini of the different mRNAs map 29, 31 and 53 nucleotides upstream from their common initiation codon. The 5′ ends of lysozyme-specific nuclear RNAs map at positions similar to that of the mRNA. AT-rich regions and sequences similar to the E. coli RNA polymerase recognition sequence are found around 30 and 70 nucleotides upstream from each of these 5′ termini. The AT-rich regions differ, however, from the canonical Goldberg-Hogness box in that they do not contain the extremely conserved TATA sequence motif. Sequence comparison at the 5′ end of the lysozyme, conalbumin and ovalbumin genes reveals only one region of partial homology, 140 nucleotides upstream from the mRNA start sites.     

The role of the chaetognath Sagitta gazellae in the vertical carbon flux of the Southern Ocean

Chaetognaths are among the most abundant predators in the Southern Ocean and are potentially important components in the biological carbon pump due to the production of large, fast-sinking fecal pellets. In situ S. gazellae abundance, fecal pellet production, sinking rates, carbon content, and vertical carbon fluxes were measured at the Lazarev Sea between December 2005 and January 2006. Sagitta gazellae produce fecal pellets that sink at speeds of 33–600 m day⁻¹ and have carbon contents of 0.01–0.8 mg C pellet⁻¹. Vertical carbon flux was later compared with the total carbon flux measured at 360 m depth at the study area. Rough estimates using published seasonal abundance of S. gazellae indicate that, at 360 m depth in the Lazarev Sea, this specie may contribute 12 and 5% of the total vertical carbon flux in winter (ice-covered) and summer (ice-free), respectively. Thus, the role of chaetognaths in the downward transport of organic matter may be far more important than previously thought.     

Plant virus infection development as affected by heavy metal stress

The work was focused on the investigation of possible dependencies between the development of viral infection in plants and the presence of high heavy metal concentrations in soil. Field experiments have been conducted in order to study the development of systemic tobacco mosaic virus (TMV) infection in Lycopersicon esculentum L. cv. Miliana plants under effect of separate salts of heavy metals Cu, Zn and Pb deposited in soil. As it is shown, simultaneous effect of viral infection and heavy metals in tenfold maximum permissible concentration leads to decrease of total chlorophyll content in experiment plants mainly due to the degradation of chlorophyll a. The reduction of chlorophyll concentration under the combined influence of both stress factors was more serious comparing to the separate effect of every single factor. Plants' treatment with toxic concentrations of lead and zinc leaded to slight delay in the development of systemic TMV infection together with more than twofold increase of virus content in plants that may be an evidence of synergism between these heavy metal's and virus' effects. Contrary, copper although decreased total chlorophyll content but showed protective properties and significantly reduced amount of virus in plants.     

The effect of past changes in inter‐annual temperature variability on tree distribution limits

Aim The northern limits of temperate broadleaved species in Fennoscanndia are controlled by their requirements for summer warmth for successful regeneration and growth as well as by the detrimental effects of winter cold on plant tissue. However, occurrences of meteorological conditions with detrimental effects on individual species are rare events rather than a reflection of average conditions. We explore the effect of changes in inter‐annual temperature variability on the abundances of the tree species Tilia cordata, Quercus robur and Ulmus glabra near their distribution limits using a process‐based model of ecosystem dynamics. Location A site in central Sweden and a site in southern Finland were used as examples for the ecotone between boreal and temperate forests in Fennoscandia. The Finnish site was selected because of the availability of varve‐thickness data. Methods The dynamic vegetation model LPJ‐GUESS was run with four scenarios of inter‐annual temperature forcing for the last 10,000 years. In one scenario the variability in the thickness of summer and winter varves from the annually laminated lake in Finland was used as a proxy for past inter‐annual temperature variability. Two scenarios were devised to explore systematically the effect of stepwise changes in the variance and shape parameter of a probability distribution. All variability scenarios were run both with and without the long‐term trend in Holocene temperature change predicted by an atmospheric general circulation model. Results Directional changes in inter‐annual temperature variability have significant effects on simulated tree distribution limits through time. Variations in inter‐annual temperature variability alone are shown to alter vegetation composition by magnitudes similar to the magnitude of changes driven by variation in mean temperatures. Main conclusions The varve data indicate that inter‐annual climate variability has changed in the past. The model results show that past changes in species abundance can be explained by changes in the inter‐annual variability of climate parameters as well as by mean climate. Because inter‐annual climatic variability is predicted to change in the future, this component of climate change should be taken into account both when making projections of future plant distributions and when interpreting vegetation history.     

Some model based considerations on observing generation times for communicable diseases

The generation time of an infectious disease is usually defined as the time from the moment one person becomes infected until that person infects another person. The concept is similar to “generation gap” in demography, with new infections replacing births in a population. Originally applied to diseases such as measles where at least the first generations are clearly discernible, the concept has recently been extended to other diseases, such as influenza, where time order of infections is usually much less apparent.By formulating the relevant statistical questions within a simple yet basic mathematical model for infection spread, it is possible to derive theoretical properties of observations in various situations e.g. in “isolation”, in households, or during large outbreaks. In each case, it is shown that the sampling distribution of observations depends on a number of factors, usually not considered in the literature and that must be taken into account in order to achieve unbiased inference about the generation time distribution. Some implications of these findings for statistical inference methods in epidemic spread models are discussed.     

Novel Binding of the Mitotic Regulator TPX2 (Target Protein for Xenopus Kinesin-like Protein 2) to Importin-��

Several aspects of mitotic spindle assembly are orchestrated by the Ran GTPase through its modulation of the interaction between spindle assembly factors and importin-α. One such factor is TPX2 that promotes microtubule assembly in the vicinity of chromosomes. TPX2 is inhibited when bound to importin-α, which occurs when the latter is bound to importin-β. The importin-α:β interaction is disrupted by the high RanGTP concentration near the chromosomes, releasing TPX2. In more distal regions, where Ran is predominantly GDP-bound, TPX2 remains bound to importin-α and so is inhibited. Here we use a combination of structural and biochemical methods to define the basis for TPX2 binding to importin-α. A 2.2 Å resolution crystal structure shows that the primary nuclear localization signal (²⁸⁴KRKH²⁸⁷) of TPX2, which has been shown to be crucial for inhibition, binds to the minor NLS-binding site on importin-α. This atypical interaction pattern was confirmed using complementary binding studies that employed importin-α variants in which binding to either the major or minor NLS-binding site was impaired, together with competition assays using the SV40 monopartite NLS that binds primarily to the major site. The different way in which TPX2 binds to importin-α could account for much of the selectivity necessary during mitosis because this would reduce the competition for binding to importin-α from other NLS-containing proteins.     

The use of modelling and simulation approach in reconstructing past landscapes from fossil pollen data: a review and results from the POLLANDCAL network

Information on past land cover in terms of absolute areas of different landscape units (forest, open land, pasture land, cultivated land, etc.) at local to regional scales is needed to test hypotheses and answer questions related to climate change (e.g. feedbacks effects of land-cover change), archaeological research, and nature conservancy (e.g. management strategy). The palaeoecological technique best suited to achieve quantitative reconstruction of past vegetation is pollen analysis. A simulation approach developed by Sugita (the computer model POLLSCAPE) which uses models based on the theory of pollen analysis is presented together with examples of application. POLLSCAPE has been adopted as the central tool for POLLANDCAL (POLlen/LANdscape CALibration), an international research network focusing on this topic. The theory behind models of the pollen–vegetation relationship and POLLSCAPE is reviewed. The two model outputs which receive greatest attention in this paper are the relevant source area of pollen (RSAP) and pollen loading in mires and lakes. Six examples of application of POLLSCAPE are presented, each of which explores a possible use of the POLLANDCAL tools and a means of validating or evaluating the models with empirical data. The landscape and vegetation factors influencing the size of the RSAP, the importance of pollen productivity estimates (PPEs) for the model outputs, the detection of small and rare patches of plant taxa in pollen records, and quantitative reconstructions of past vegetation and landscapes are discussed on the basis of these examples. The simulation approach is seen to be useful both for exploring different vegetation/landscape scenarios and for refuting hypotheses.