Roles for Ca2+ mobilization and its regulation in mast cell functions: recent progress

Ca(2+)mobilization in response to cross-linking of IgE bound to its high affinity receptor, FcεRI, on mast cells is central to immune allergic responses. Stimulated tyrosine phosphorylation caused by this cross-linking activates store-operated Ca(2+)entry that results in sustained Ca(2+)oscillations dependent on Rho family GTPases and phosphoinositide synthesis. Coupling of the endoplasmic reticulum (ER) Ca(2+)sensor, stromal interaction molecule 1 (STIM1), to the Ca(2+)-selective channel, Orai1, is regulated by these elements and depends on membrane organization, both at the plasma membrane and at the ER. Mitochondria also contribute to the regulation of Ca(2+)mobilization, and we describe recent evidence that the ER membrane protein vesicle-associated membrane protein-associated protein (VAP) plays a significant role in the coupling between ER and mitochondria in this process. In addition to granule exocytosis, Ca(2+)mobilization in these cells also contributes to stimulated outward trafficking of recycling endosomes and to antigen-stimulated chemotaxis, and it is pathologically regulated by protozoan parasitic invasion

Allelomimesis as universal clustering mechanism for complex adaptive systems

Animal and human clusters are complex adaptive systems and many are organized in cluster sizes $s$ that obey the frequency-distribution $D(s)\propto s^{-\tau}$. Exponent $\tau$ describes the relative abundance of the cluster sizes in a given system. Data analyses have revealed that real-world clusters exhibit a broad spectrum of $\tau$-values, $0.7\textrm{(tuna fish schools)}\leq\tau\leq 2.95\textrm{(galaxies)}$. We show that allelomimesis is a fundamental mechanism for adaptation that accurately explains why a broad spectrum of $\tau$-values is observed in animate, human and inanimate cluster systems. Previous mathematical models could not account for the phenomenon. They are hampered by details and apply only to specific systems such as cities, business firms or gene family sizes. Allelomimesis is the tendency of an individual to imitate the actions of its neighbors and two cluster systems yield different $\tau$ values if their component agents display different allelomimetic tendencies. We demonstrate that allelomimetic adaptation are of three general types: blind copying, information-use copying, and non-copying. Allelomimetic adaptation also points to the existence of a stable cluster size consisting of three interacting individuals.Comment: 8 pages, 5 figures, 2 table

Stereochemical control of the DNA binding affinity, sequence specificity, and orientation preference of chiral hairpin polyamides in the minor groove

Three-ring polyamides containing pyrrole (Py) and imidazole (Im) amino acids covalently coupled by γ-aminobutyric acid (γ) form six-ring hairpins that recognize five-base-pair sequences in the minor groove of DNA. Selective chiral substitution of the ``gamma-turn'' enhances the properties of polyamide hairpins with regard to DNA affinity and sequence specificity. Polyamides of core sequence composition ImPyPy-γ-PyPyPy which differ by selective stereochemical substitution of the prochiral alpha-position in the γ-turn were prepared. The DNA binding properties of two enantiomeric polyamides were analyzed by footprinting and affinity cleavage on a DNA fragment containing two match sites (5'-TGTTA-3' and 5'-ACATT-3') and one 5'-TGTCA-3' mismatch site. Quantitative footprint titrations demonstrate that replacement of γ-aminobutyric acid by (R)-2,4-diaminobutyric acid enhances DNA binding affinity for the 5'-TGTTA-3' match site 13-fold (K_a = 3.8 x 10^9 M-1). The enhanced affinity is achieved without a compromise in sequence selectivity, which in fact increases and is found to be 100-fold higher relative to binding at a single base pair mismatch sequence, 5'-TGTCA-3'. An (S)-2,4-diaminobutyric acid linked hairpin binds with 170-fold reduced affinity relative to the R-enantiomer and only 5-fold sequence specificity versus a 5'-ACATT-3' reversed orientation site. These effects are modulated by acetylation of the chiral amine substituents. This study identifies structural elements which should facilitate the design of new hairpin polyamides with improved DNA binding affinity, sequence specificity, and orientational selectivity

Inhibition of RNA polymerase II transcription in human cells by synthetic DNA-binding ligands

Sequence-specific DNA-binding small molecules that can permeate human cells potentially could regulate transcription of specific genes. Multiple cellular DNA-binding transcription factors are required by HIV type 1 for RNA synthesis. Two pyrrole-imidazole polyamides were designed to bind DNA sequences immediately adjacent to binding sites for the transcription factors Ets-l, lymphoid-enhancer binding factor 1, and TATA-box binding protein. These synthetic ligands specifically inhibit DNA-binding of each transcription factor and HIV type 1 transcription in cell-free assays. When used in combination, the polyamides inhibit virus replication by >99% in isolated human peripheral blood lymphocytes, with no detectable cell toxicity, The ability of small molecules to target predetermined DNA sequences located within RNA polymerase II promoters suggests a general approach for regulation of gene expression, as well as a mechanism for the inhibition of viral replication

Scaling Behaviour and Complexity of the Portevin-Le Chatelier Effect

The plastic deformation of dilute alloys is often accompanied by plastic instabilities due to dynamic strain aging and dislocation interaction. The repeated breakaway of dislocations from and their recapture by solute atoms leads to stress serrations and localized strain in the strain controlled tensile tests, known as the Portevin-Le Chatelier (PLC) effect. In this present work, we analyse the stress time series data of the observed PLC effect in the constant strain rate tensile tests on Al-2.5%Mg alloy for a wide range of strain rates at room temperature. The scaling behaviour of the PLC effect was studied using two complementary scaling analysis methods: the finite variance scaling method and the diffusion entropy analysis. From these analyses we could establish that in the entire span of strain rates, PLC effect showed Levy walk property. Moreover, the multiscale entropy analysis is carried out on the stress time series data observed during the PLC effect to quantify the complexity of the distinct spatiotemporal dynamical regimes. It is shown that for the static type C band, the entropy is very low for all the scales compared to the hopping type B and the propagating type A bands. The results are interpreted considering the time and length scales relevant to the effect.Comment: 35 pages, 6 figure

Measles vaccination and antibody response in autism spectrum disorder

OBJECTIVE: To test the hypothesis that measles vaccination was involved in the pathogenesis of autism spectrum disorders (ASD) as evidenced by signs of a persistent measles infection or abnormally persistent immune response shown by circulating measles virus or raised antibody titres in children with ASD who had been vaccinated against measles, mumps and rubella (MMR) compared with controls. DESIGN: Case-control study, community based. METHODS: A community sample of vaccinated children aged 10-12 years in the UK with ASD (n = 98) and two control groups of similar age, one with special educational needs but no ASD (n = 52) and one typically developing group (n = 90), were tested for measles virus and antibody response to measles in the serum. RESULTS: No difference was found between cases and controls for measles antibody response. There was no dose-response relationship between autism symptoms and antibody concentrations. Measles virus nucleic acid was amplified by reverse transcriptase-PCR in peripheral blood mononuclear cells from one patient with autism and two typically developing children. There was no evidence of a differential response to measles virus or the measles component of the MMR in children with ASD, with or without regression, and controls who had either one or two doses of MMR. Only one child from the control group had clinical symptoms of possible enterocolitis. CONCLUSION: No association between measles vaccination and ASD was shown

Fmoc solid phase synthesis of polyamides containing pyrrole and imidazole amino acids

Polyamides containing N-methylimidazole (Im) and N-methylpyrrole (Py) amino acids are synthetic ligands that have an affinity and specificity for DNA comparable to those of many naturally occurring DNA binding proteins. A machine-assisted Fmoc solid phase synthesis of polyamides has been optimized to afford high stepwise coupling yields (>99%). Two monomer building blocks, Fmoc-Py acid and Fmoc-Im acid, were prepared in multigram scale. Cleavage by aminolysis followed by HPLC purification affords up to 200 mg quantities of polyamide with purities and yields greater than or equal to those reported using Boc chemistry. A broader set of reaction conditions will increase the number and complexity of minor groove binding polyamides which may be prepared and help ensure compatibility with many commercially available peptide synthesizers

Aliphatic/aromatic amino acid pairings for polyamide recognition in the minor groove of DNA

Selective placement of an aliphatic β-alanine (β) residue paired side-by-side with either a pyrrole (Py) or imidazole (Im) aromatic amino acid is found to compensate for sequence composition effects for recognition of the minor groove of DNA by hairpin pyrrole−imidazole polyamides. A series of polyamides were prepared which contain pyrrole and imidazole aromatic amino acids, as well as γ-aminobutyric acid (γ) “turn” and β-alanine “spring” aliphatic amino acid residues. The binding affinities and specificities of these polyamides are regulated by the placement of paired β/β, Py/β, and Im/β residues. Quantitative footprint titrations demonstrate that replacing two Py/Py pairings in a 12-ring hairpin (6-γ-6) with two Py/β pairings affords 10-fold enhanced affinity and similar sequence specificity for an 8-bp target sequence. The 6-γ-6 hairpin ImPyImPyPyPy-γ-ImPyPyPyPyPy-β-Dp, which contains six consecutive amino acid pairings, is unable to discriminate a single-base-pair mismatch site 5‘-TGTTAACA-3‘ from a 5‘-TGTGAACA-3‘ match site. The hairpin polyamide Im-β-ImPyPyPy-γ-ImPyPyPy-β-Py-β-Dp binds to the 8-bp match sequence 5‘-TGTGAACA-3‘ with an equilibrium association constant of Ka = 2.4 × 1010 M-1 and ≥48-fold specificity versus the 5‘-TGTTAACA-3‘ single-base-pair mismatch site. Modeling indicates that the β-alanine residue relaxes ligand curvature, providing for optimal hydrogen bond formation between the floor of the minor groove and both Im residues within the Im-β-Im polyamide subunit. This observation provided the basis for design of a hairpin polyamide, Im-β-ImPy-γ-Im-β-ImPy-β-Dp, which incorporates Im/β pairings to recognize a “problematic” 5‘-GCGC-3‘ sequence at subnanomolar concentrations. These results identify Im/β and β/Im pairings that respectively discriminate G·C and C·G from A·T/T·A as well as Py/β and β/Py pairings that discriminate A·T/T·A from G·C/C·G. These aliphatic/aromatic amino acid pairings will facilitate the design of hairpin polyamides which recognize both a larger binding site size as well as a more diverse sequence repertoire

Ovarian hormones and pituitary responsiveness to gonadotrophin releasing hormone in mice

In the present study the role of progesterone and oestradiol in modulating the responsiveness at the anterior pituitary gland to gonadotrophin releasing hormone (GN-RH) was investigated by measuring the release of luteinizing hormone (LH) in ovariectomised mice which had been pretreated with these steroids.A significant release of LH was seen in the animals receiving GN-RH. Pretreatment with oestrogen and progesterone depressed LH levels in the animals which did not receive GN-RH, and resulted in a larger release of LH in the animals receiving GN-RH in comparison with the control group pretreated with oil