Inter-DNA Attraction Mediated by Divalent Counterions

Can nonspecifically bound divalent counterions induce attraction between DNA strands? Here, we present experimental evidence demonstrating attraction between short DNA strands mediated by Mg2 ions. Solution small angle x-ray scattering data collected as a function of DNA concentration enable model independent extraction of the second virial coefficient. As the [Mg2] increases, this coefficient turns from positive to negative reflecting the transition from repulsive to attractive inter-DNA interaction. This surprising observation is corroborated by independent light scattering experiments. The dependence of the observed attraction on experimental parameters including DNA length provides valuable clues to its origin

Measuring Inter-DNA Potentials in Solution

Interactions between short strands of DNA can be tuned from repulsive to attractive by varying solution conditions and have been quantified using small angle x-ray scattering techniques. The effective DNA interaction charge was extracted by fitting the scattering profiles with the generalized one-component method and inter-DNA Yukawa pair potentials. A significant charge is measured at low to moderate monovalent counterion concentrations, resulting in strong inter-DNA repulsion. The charge and repulsion diminish rapidly upon the addition of divalent counterions. An intriguing short range attraction is observed at surprisingly low divalent cation concentrations, ~16 mM Mg2+. Quantitative measurements of inter- DNA potentials are essential for improving models of fundamental interactions in biological systems

Mono- and Trivalent Ions around DNA: A Small-Angle Scattering Study of Competition and Interactions

The presence of small numbers of multivalent ions in DNA-containing solutions results in strong attractive forces between DNA strands. Despite the biological importance of this interaction, e.g., DNA condensation, its physical origin remains elusive.Wecarried out a series of experiments to probe interactions between short DNA strands as small numbers of trivalent ions are included in a solution containing DNA and monovalent ions. Using resonant (anomalous) and nonresonant small angle x-ray scattering, we coordinated measurements of the number and distribution of each ion species around the DNA with the onset of attractive forces between DNA strands. DNA-DNA interactions occur as the number of trivalent ions increases. Surprisingly good agreement is found between data and size-corrected numerical Poisson-Boltzmann predictions of ion competition for non- and weakly interacting DNAs. We also obtained an estimate for the minimum number of trivalent ions needed to initiate DNA-DNA attraction

Focusing Capillary Optics for Use in Solution Small-Angle X-Ray Scattering

Measurements of the global conformation of macromolecules can be carried out using small-angle X-ray scattering (SAXS). Glass focusing capillaries, manufactured at the Cornell High Energy Synchrotron Source (CHESS), have been successfully employed for SAXS measurements on the heme protein cytochrome c. These capillaries provide high X-ray flux into a spot size of tens of micrometres, permitting short exposures of small-volume samples. Such a capability is ideal for use in conjunction with microfluidic mixers, where time resolution may be determined by beam size and sample volumes are kept small to facilitate mixing and conserve material

Spatial Distribution of Competing Ions around DNA in Solution

The competition of monovalent and divalent cations for proximity to negatively charged DNA is of biological importance and can provide strong constraints for theoretical treatments of polyelectrolytes. Resonant x-ray scattering experiments have allowed us to monitor the number and distribution of each cation in a mixed ion cloud around DNA. These measurements provide experimental evidence to support a general theoretical prediction: the normalized distribution of each ion around polyelectrolytes remains constant when ions are mixed at different ratios. In addition, the amplitudes of the scattering signals throughout the competition provide a measurement of the surface concentration parameter that predicts the competition behavior of these cations. The data suggest that ion size needs to be taken into account in applying Poisson-Boltzmann treatments to polyelectrolytes such as DNA

Multinational Web Uses and Gratifications: Measuring the Social Impact of Online Community Participation Across National Boundaries

This paper describes the rationale and findings from a multinational study of online uses and gratifications conducted in theUnited States,Korea, and theNetherlandsin spring 2003. Survey questions developed in three languages by native speaking researchers was presented to approximately 400 respondents in each country via the Web. Web uses and gratifications were analyzed cross-nationally in a comparative fashion focusing on involvement in different types of on-line communities. Findings indicate that demographic characteristics, cultural values, and Internet connection type emerged as critical factors that explain why the same technology is adopted differently

Differences in the Curing of [PSI+] Prion by Various Methods of Hsp104 Inactivation

[PSI+] yeast, containing the misfolded amyloid conformation of Sup35 prion, is cured by inactivation of Hsp104. There has been controversy as to whether inactivation of Hsp104 by guanidine treatment or by overexpression of the dominant negative Hsp104 mutant, Hsp104-2KT, cures [PSI+] by the same mechanism– inhibition of the severing of the prion seeds. Using live cell imaging of Sup35-GFP, overexpression of Hsp104-2KT caused the foci to increase in size, then decrease in number, and finally disappear when the cells were cured, similar to that observed in cells cured by depletion of Hsp104. In contrast, guanidine initially caused an increase in foci size but then the foci disappeared before the cells were cured. By starving the yeast to make the foci visible in cells grown with guanidine, the number of cells with foci was found to correlate exactly with the number of [PSI+] cells, regardless of the curing method. Therefore, the fluorescent foci are the prion seeds required for maintenance of [PSI+] and inactivation of Hsp104 cures [PSI+] by preventing severing of the prion seeds. During curing with guanidine, the reduction in seed size is an Hsp104-dependent effect that cannot be explained by limited severing of the seeds. Instead, in the presence of guanidine, Hsp104 retains an activity that trims or reduces the size of the prion seeds by releasing Sup35 molecules that are unable to form new prion seeds. This Hsp104 activity may also occur in propagating yeast

Huntingtin Fragments and SOD1 Mutants Form Soluble Oligomers in the Cell

Diffusion coefficients of huntingtin (Htt) fragments and SOD1 mutants expressed in cells were measured using fluorescence correlation spectroscopy. The diffusion mobilities of both non-pathological Htt fragments (25 polyQs) and pathological Htt fragments (103 polyQs) were much slower than expected for monomers suggesting that they oligomerize. The mobility of these fragments was unaffected by duration of expression or by over-expression of Hsp70 and Hsp40. However in cells with HttQ103 inclusions, diffusion measurements showed that the residual cytosolic HttQ103 was monomeric. These results suggest that both non-pathological and pathological Htt fragments form soluble oligomers in the cytosol with the properties of the oligomers determining whether they cause pathology. SOD1 with point mutations (A4V, G37R, and G85R) also had slower diffusional mobility than the wild-type protein whose mobility was consistent with that of a dimer. However, the decrease in mobility of the different SOD1 mutantsdid not correlate with their known pathology. Therefore, while soluble oligomers always seem to be present under conditions where cell pathology occurs, the presence of the oligomers, in itself, does not determine the extent of neuropathology