Role of c-Jun in cellular sensitivity to the microtubule inhibitor vinblastine

The role of c-Jun in the apoptotic response of cells to the microtubule inhibitor vinblastine was investigated using fibroblasts lacking or overexpressing c-Jun. c-Jun null cells were found to be more sensitive than wild-type cells at low (1-3 nM) concentrations of vinblastine, but showed essentially identical apoptotic responses as wild-type cells at a higher concentration of 10nM. In contrast, c-Jun overexpressing cells were highly vinblastine-resistant, with an IC50 of 12-fold greater than wild-type cells. The fate of cells exposed to lethal concentrations of vinblastine was examined by propidium iodide staining and flow cytometry. All cell types appeared to undergo mitotic arrest prior to apoptosis. Apoptosis of wild-type cells was associated with significant DNA re-replication. In contrast, DNA re-replication was much less prominent in vinblastine-treated c-Jun null cells and absent during apoptosis of c-Jun overexpressing cells. These results suggest that c-Jun plays a key role in the cellular sensitivity to vinblastine. In addition, c-Jun appears to regulate the pathway to cell death following mitotic arrest.     

The SUN family of Saccharomyces cerevisiae: the double knock-out of UTH1 and SIM1 promotes defects in nucleus migration and increased drug sensitivity

UTH1 and SIM1 are two of four 'SUN' genes (SIM1, UTH1, NCA3 and SUN4/SCW3) whose products are involved in different cellular processes such as DNA replication, lifespan, mitochondrial biogenesis or cell septation. UTH1 or SIM1 inactivation did not affect cell growth, shape or nuclear migration, whereas the double null mutant presented phenotypes of numerous binucleate cells and benomyl sensitivity, suggesting that microtubule function could be altered; the uth1Deltasim1Delta strain also presented defects which could be related to the Ras/cAMP pathway: pet phenotype, heat shock sensitivity, inability to store glycogen, sensitivity to starvation and failure of spores to germinate. These observations suggested that Uth1p could be involved as a connection step between pathways controlling growth and those controlling division.     

Temperature Sensitivity of the Tonoplast Ca2+-Activated K+ Channel in Chara: The Influence of Reversing the Sign of Membrane Potential

A detailed temperature dependence study of a well-defined plant ion channel, the Ca²⁺-activated K⁺ channel of Chara corallina, was performed over the temperature range of their habitats, 5–36°C, at 1°C resolution. The temperature dependence of the channel unitary conductance at 50 mV shows discontinuities at 15 and 30°C. These temperatures limit the range within which ion diffusion is characterized by the lowest activation energy (E _a = 8.0 ± 1.6 kJ/mol) as compared to the regions below 15°C and above 30°C. Upon reversing membrane voltage polarity from 50 to −50 mV the pattern of temperature dependence switched from discontinuous to linear with E _a = 13.6 ± 0.5 kJ/mol. The temperature dependence of the effective number of open channels at 50 mV showed a decrease with increasing temperature, with a local minimum at 28°C. The mean open time exhibited a similar behavior. Changing the sign of membrane potential from 50 to −50 mV abolished the minima in both temperature dependencies. These data are discussed in the light of higher order phase transitions of the Characean membrane lipids and corresponding change in the lipid-protein interaction, and their modulation by transmembrane voltage. Received: 14 June 2000/Revised: 20 September 2000     

Chaos and population control of insect outbreaks

We used small perturbations in adult numbers to control large fluctuations in the chaotic demographic dynamics of laboratory populations of the flour beetle Tribolium castaneum . A nonlinear mathematical model was used to identify a sensitive region of phase space where the addition of a few adult insects would result in a dampening of the life stage fluctuations. Three experimental treatments were applied: one in which perturbations were made whenever the populations were inside the sensitive region ("in-box treatment"), another where perturbations were made whenever the populations were outside the sensitive region ("out-box treatment"), and an unperturbed control. The in-box treatment caused a stabilization of insect densities at numbers well below the peak values exhibited by the out-box and control populations. This study demonstrates how small perturbations can be used to influence the chaotic dynamics of an ecological system.     

Wavelength- and Time-Dependence of Potentiometric Non-linear Optical Signals from Styryl Dyes

Second harmonic generation (SHG) imaging microscopy is an important emerging technique for biological research, complementing existing one- and two-photon fluorescence (2PF) methods. A non-linear phenomenon employing light from mode-locked Ti:sapphire or fiber-based lasers, SHG results in intrinsic optical sectioning without the need for a confocal aperture. Furthermore, as a second-order process SHG is confined to loci lacking a center of symmetry, a constraint that is readily satisfied by lipid membranes with only one leaflet stained by a dye. Of particular interest is “resonance-enhanced” SHG from styryl dyes in cellular membranes and the possibility that SHG is sensitive to transmembrane potential. We have previously confirmed this, using simultaneous voltage-clamping and non-linear imaging of cells to find that SHG is up to four times more sensitive to potential than fluorescence. In this work, we have extended these results in two directions. First, with a range of wavelengths available from a mode-locked Ti:sapphire laser and a fiber-based laser, we have more fully investigated SHG and 2PF voltage-sensitivity from ANEP and ASTAP chromophores, obtaining SHG sensitivity spectra that are consistent with resonance enhancements. Second, we have modified our system to coordinate the application of voltage-clamp steps with non-linear image acquisition to more precisely characterize the time dependence of SHG and 2PF voltage sensitivity, finding that, at least for some dyes, SHG responds more slowly than fluorescence to changes in transmembrane potential.     

Temperature Broadening of LH2 Absorption in Glycerol Solution

In order to determine the relationship between the pigment–protein and the pigment–pigment interactions, the measurements of absorption spectra of the peripheral light-harvesting complex LH2 from the purple bacteria Rhodobacter sphaeroides solvated in glycerol/buffer solution were carried out in a wide temperature range, from 4 to 250 K. The SDFs used for simulating the temperature dependence of B800 and B850 bands were determined in a parametric form. To fit experimental spectra the overall exciton–phonon coupling had to be assumed to be weak for B850 (λ/2V ≈ 0.3, where λ is the reorganization energy and V is the nearest-neighbor dipole–dipole coupling for bacteriochlorophylls). At physiological temperatures the intermediate nuclear bath dynamics compares with the magnitude of energy gap fluctuations. Slower dynamics with κ ≈ 0.39, where κ is the ratio of the nuclear relaxation rate and the line width parameter, determines the spectral shape of B850 whilst faster modulations characterize B800 (κ ≈2.39). The static disorder for the B800 band is relatively high with the characteristic value of the inhomogeneous bandwidth Γ^inh ≈120 cm⁻¹, while for the B850 band this value is almost equal to the dipole–dipole coupling strength (Γ^inh ≈360 cm⁻¹). It has been found that the LH2 absorption spectrum is likely to be influenced by the temperature dependence of the dielectric constant of the solution in the high temperature range, when the glycerol/buffer solution is in the liquid state.     

Spectral Study of Interactions of 4,8,4′-Trimethylpsoralen and 4,4′-Dimethylangelicin Dyes with DNA

Absorption and fluorescence spectra for six new synthetic dyes of 4,8,4′-trimethylpsoralen and 4,4′-dimethylangelicin derivatives containing various terminal substituents at 5′-position have been investigated in different environments using a wide range of the DNA/ligand concentrations. Various spectral and binding characteristics of the DNA-ligand systems have been determined. General principles characterizing mechanisms responsible for changes in the fluorescent properties of nucleotide-specific dyes have been proposed; they take into consideration chemical structure of the dyes, properties of the environment, and degree of sorption on substrate.__________Translated from Biokhimiya, Vol. 70, No. 7, 2005, pp. 995–1007.Original Russian Text Copyright © 2005 by Sibirtsev, Tolmachev, Kovaleva, Garabadzhiu, Traven.     

Rapid response to habitat restoration by the perennial Primula veris as revealed by demographic monitoring

The demography of Primula veris, a typical species of the species-rich Mesobromion grasslands, was investigated at two contrasting habitats in Eastern Belgium. Both a forested site and a clear-cut parcel were part of formerly larger calcareous grassland areas. By monitoring the demographic response of the target species, this study attempts to clarify the differences in fecundity, growth and survival between a restored and degraded calcareous grassland site. A study of plant traits showed a decrease in number of flowers, inflorescence stalks and plant size under a closing tree canopy. We surveyed individuals in permanent plots between 1999 and 2001. At the forested site, a first sign of decline included lower proportions of flowering individuals and afterwards an increase in mortality was found. Alternatively, removing canopy resulted in an immediate flowering response and both increased growth and seedling recruitment the year after. At both sites, survival rates strongly depended on the initial state, location and year. Projection matrix analysis revealed large differences in modelled population growth rates between sites and years. Under closing canopy the species showed only slow population decline, but the decrease was larger in the last survey year when higher mortality affected the number of reproductive individuals. Transitions between stages with the highest impact on population growth rate were identified by elasticity analysis. If calcareous grassland is forested, survival of reproductive adults is predicted to be very important for conservation of P. veris populations. However, seedling recruitment needs to be raised to guarantee long-term persistence of the populations. On the other hand, clear-felling of a site for restoration of species-rich calcareous grasslands may result in a rapid recovery of certain species.     

Testing hypotheses about ecological specialization using phylogenetic trees

It is often assumed that ecological specialization represents an evolutionary "dead-end" that limits further evolution. Maximum-likelihood (ML) analyses on phylogenies for 15 groups of phytophagous insects revealed that high transition rates both to and from specialization occurred, but that the mean ratio of rates was significantly biased toward a higher rate to specialization. Here we explore the consequences of the fact that transition rates inferred by ML are affected not only by the distribution, but also by the frequency, of character states. Higher rates to the more common state were inferred in the analyses of Nosil (2002), in similar studies published since 2002, and in a small set of simulations. Thus, the ratio of the rate toward versus away from specialization was strongly, positively correlated with the proportion of specialist species at the tips of the phylogeny and whether transitions away from specialization occur remains unclear. Here we reexamine these data using methods that do not rely on directly comparing transition rates. Maximum-likelihood analyses show that models with no transitions in one direction (e.g., irreversible evolution as predicted by the "specialist as dead end" framework) are usually strongly rejected, independent of the proportion of specialists at the tips. Ancestral state reconstruction revealed two instances where generalists were unambiguously derived from specialists. Transition rates would need to biased 100-fold and 5000-fold toward specialization to reconstruct a history where these shifts from specialization toward generalization do not occur. The general conclusions of Nosil (2002) appear to hold; transitions in either direction likely occur such that specialization does not always limit further evolution. Most generally, inferences regarding character evolution can be strengthened by comparing models of character change and examining ancestor states, rather than only comparing parameter values.