Self-organized defensive behavior in honeybees

We investigated the defensive behavior of honeybees under controlled experimental conditions. During an attack on two identical targets, the spatial distribution of stings varied as a function of the total number of stings, evincing the classic “pitchfork bifurcation” phenomenon of nonlinear dynamics. The experimental results support a model of defensive behavior based on a self-organizing mechanism. The model helps to explain several of the characteristic features of the honeybee defensive response: (i) the ability of the colony to localize and focus its attack, (ii) the strong variability between different hives in the intensity of attack, as well as (iii) the variability observed within the same hive, and (iv) the ability of the colony to amplify small differences between the targets.

Self-organized phase transitions in neural networks as a neural mechanism of information processing.

Transitions between dynamically stable activity patterns imposed on an associative neural network are shown to be induced by self-organized infinitesimal changes in synaptic connection strength and to be a kind of phase transition. A key event for the neural process of information processing in a population coding scheme is transition between the activity patterns encoding usual entities. We propose that the infinitesimal and short-term synaptic changes based on the Hebbian learning rule are the driving force for the transition. The phase transition between the following two dynamical stable states is studied in detail, the state where the firing pattern is changed temporally so as to itinerate among several patterns and the state where the firing pattern is fixed to one of several patterns. The phase transition from the pattern itinerant state to a pattern fixed state may be induced by the Hebbian learning process under a weak input relevant to the fixed pattern. The reverse transition may be induced by the Hebbian unlearning process without input. The former transition is considered as recognition of the input stimulus, while the latter is considered as clearing of the used input data to get ready for new input. To ensure that information processing based on the phase transition can be made by the infinitesimal and short-term synaptic changes, it is absolutely necessary that the network always stays near the critical state corresponding to the phase transition point.

In situ atomic force microscopy study of Alzheimer’s β-amyloid peptide on different substrates: New insights into mechanism of β-sheet formation

We have applied in situ atomic force microscopy to directly observe the aggregation of Alzheimer’s β-amyloid peptide (Aβ) in contact with two model solid surfaces: hydrophilic mica and hydrophobic graphite. The time course of aggregation was followed by continuous imaging of surfaces remaining in contact with 10–500 μM solutions of Aβ in PBS (pH 7.4). Visualization of fragile nanoscale aggregates of Aβ was made possible by the application of a tapping mode of imaging, which minimizes the lateral forces between the probe tip and the sample. The size and the shape of Aβ aggregates, as well as the kinetics of their formation, exhibited pronounced dependence on the physicochemical nature of the surface. On hydrophilic mica, Aβ formed particulate, pseudomicellar aggregates, which at higher Aβ concentration had the tendency to form linear assemblies, reminiscent of protofibrillar species described recently in the literature. In contrast, on hydrophobic graphite Aβ formed uniform, elongated sheets. The dimensions of those sheets were consistent with the dimensions of β-sheets with extended peptide chains perpendicular to the long axis of the aggregate. The sheets of Aβ were oriented along three directions at 120° to each other, resembling the crystallographic symmetry of a graphite surface. Such substrate-templated self-assembly may be the distinguishing feature of β-sheets in comparison with α-helices. These studies show that in situ atomic force microscopy enables direct assessment of amyloid aggregation in physiological fluids and suggest that Aβ fibril formation may be driven by interactions at the interface of aqueous solutions and hydrophobic substrates, as occurs in membranes and lipoprotein particles in vivo.

Self-antigen does not accelerate immature B cell apoptosis, but stimulates receptor editing as a consequence of developmental arrest

In pre-B lymphocytes, productive rearrangement of Ig light chain genes allows assembly of the B cell receptor (BCR), which selectively promotes further developmental maturation through poorly defined transmembrane signaling events. Using a novel in vitro system to study immune tolerance during development, we find that BCR reactivity to auto-antigen blocks this positive selection, preventing down-regulation of light chain gene recombination and promoting secondary light chain gene rearrangements that often alter BCR specificity, a process called receptor editing. Under these experimental conditions, self-antigen induces secondary light chain gene rearrangements in at least two-thirds of autoreactive immature B cells, but fails to accelerate cell death at this stage. These data suggest that in these cells the mechanism of immune tolerance is receptor selection rather than clonal selection.

Striking sequence similarity in inter- and intra-specific comparisons of class I SLG alleles from Brassica oleracea and Brassica campestris: Implications for the evolution and recognition mechanism

Self-incompatibility in Brassica is controlled by a single multi-allelic locus (S locus), which contains at least two highly polymorphic genes expressed in the stigma: an S glycoprotein gene (SLG) and an S receptor kinase gene (SRK). The putative ligand-binding domain of SRK exhibits high homology to the secretory protein SLG, and it is believed that SLG and SRK form an active receptor kinase complex with a self-pollen ligand, which leads to the rejection of self-pollen. Here, we report 31 novel SLG sequences of Brassica oleracea and Brassica campestris. Sequence comparisons of a large number of SLG alleles and SLG-related genes revealed the following points. (i) The striking sequence similarity observed in an inter-specific comparison (95.6% identity between SLG14 of B. oleracea and SLG25 of B. campestris in deduced amino acid sequence) suggests that SLG diversification predates speciation. (ii) A perfect match of the sequences in hypervariable regions, which are thought to determine S specificity in an intra-specific comparison (SLG8 and SLG46 of B. campestris) and the observation that the hypervariable regions of SLG and SRK of the same S haplotype were not necessarily highly similar suggests that SLG and SRK bind different sites of the pollen ligand and that they together determine S specificity. (iii) Comparison of the hypervariable regions of SLG alleles suggests that intragenic recombination, together with point mutations, has contributed to the generation of the high level of sequence variation in SLG alleles. Models for the evolution of SLG/SRK are presented.

Mechanism of protein remodeling by ClpA chaperone

ClpA, a newly discovered ATP-dependent molecular chaperone, remodels bacteriophage P1 RepA dimers into monomers, thereby activating the latent specific DNA binding activity of RepA. We investigated the mechanism of the chaperone activity of ClpA by dissociating the reaction into several steps and determining the role of nucleotide in each step. In the presence of ATP or a nonhydrolyzable ATP analog, the initial step is the self-assembly of ClpA and its association with inactive RepA dimers. ClpA-RepA complexes form rapidly and at 0°C but are relatively unstable. The next step is the conversion of unstable ClpA-RepA complexes into stable complexes in a time- and temperature-dependent reaction. The transition to stable ClpA-RepA complexes requires binding of ATP, but not ATP hydrolysis, because nonhydrolyzable ATP analogs satisfy the nucleotide requirement. The stable complexes contain approximately 1 mol of RepA dimer per mol of ClpA hexamer and are committed to activating RepA. In the last step of the reaction, active RepA is released upon exchange of ATP with the nonhydrolyzable ATP analog and ATP hydrolysis. Importantly, we discovered that one cycle of RepA binding to ClpA followed by ATP-dependent release is sufficient to convert inactive RepA to its active form.

Inhibition of interferon γ induced interleukin 12 production: A potential mechanism for the anti-inflammatory activities of tumor necrosis factor

Inflammation is associated with production of cytokines and chemokines that recruit and activate inflammatory cells. Interleukin (IL) 12 produced by macrophages in response to various stimuli is a potent inducer of interferon (IFN) γ production. IFN-γ, in turn, markedly enhances IL-12 production. Although the immune response is typically self-limiting, the mechanisms involved are unclear. We demonstrate that IFN-γ inhibits production of chemokines (macrophage inflammatory proteins MIP-1α and MIP-1β). Furthermore, pre-exposure to tumor necrosis factor (TNF) inhibited IFN-γ priming for production of high levels of IL-12 by macrophages in vitro. Inhibition of IL-12 by TNF can be mediated by both IL-10-dependent and IL-10-independent mechanisms. To determine whether TNF inhibition of IFN-γ-induced IL-12 production contributed to the resolution of an inflammatory response in vivo, the response of TNF+/+ and TNF−/− mice injected with Corynebacterium parvum were compared. TNF−/− mice developed a delayed, but vigorous, inflammatory response leading to death, whereas TNF+/+ mice exhibited a prompt response that resolved. Serum IL-12 levels were elevated 3-fold in C. parvum-treated TNF−/− mice compared with TNF+/+ mice. Treatment with a neutralizing anti-IL-12 antibody led to resolution of the response to C. parvum in TNF−/− mice. We conclude that the role of TNF in limiting the extent and duration of inflammatory responses in vivo involves its capacity to regulate macrophage IL-12 production. IFN-γ inhibition of chemokine production and inhibition of IFN-γ-induced IL-12 production by TNF provide potential mechanisms by which these cytokines can exert anti-inflammatory/repair function(s).

Ubiquitously Expressed Dynamin-II Has a Higher Intrinsic GTPase Activity and a Greater Propensity for Self-assembly Than Neuronal Dynamin-I

To begin to understand mechanistic differences in endocytosis in neurons and nonneuronal cells, we have compared the biochemical properties of the ubiquitously expressed dynamin-II isoform with those of neuron-specific dynamin-I. Like dynamin-I, dynamin-II is specifically localized to and highly concentrated in coated pits on the plasma membrane and can assemble in vitro into rings and helical arrays. As expected, the two closely related isoforms share a similar mechanism for GTP hydrolysis: both are stimulated in vitro by self-assembly and by interaction with microtubules or the SH3 domain-containing protein, grb2. Deletion of the C-terminal proline/arginine-rich domain from either isoform abrogates self-assembly and assembly-dependent increases in GTP hydrolysis. However, dynamin-II exhibits a ∼threefold higher rate of intrinsic GTP hydrolysis and higher affinity for GTP than dynamin-I. Strikingly, the stimulated GTPase activity of dynamin-II can be >40-fold higher than dynamin-I, due principally to its greater propensity for self-assembly and the increased resistance of assembled dynamin-II to GTP-triggered disassembly. These results are consistent with the hypothesis that self-assembly is a major regulator of dynamin GTPase activity and that the intrinsic rate of GTP hydrolysis reflects a dynamic, GTP-dependent equilibrium of assembly and disassembly.

Redox state is a central modulator of the balance between self-renewal and differentiation in a dividing glial precursor cell

We have discovered that intracellular redox state appears to be a necessary and sufficient modulator of the balance between self-renewal and differentiation in dividing oligodendrocyte-type-2 astrocyte progenitor cells. The intracellular redox state of freshly isolated progenitors allows prospective isolation of cells with different self-renewal characteristics. Redox state is itself modulated by cell-extrinsic signaling molecules that alter the balance between self-renewal and differentiation: growth factors that promote self-renewal cause progenitors to become more reduced, while signaling molecules that promote differentiation cause progenitors to become more oxidized. Moreover, pharmacological antagonists of the redox effects of these cell-extrinsic signaling molecules antagonize their effects on self-renewal and differentiation, indicating that cell-extrinsic signaling molecules that modulate this balance converge on redox modulation as a critical component of their effector mechanism.

Involvement of a cytosine side chain in proton transfer in the rate-determining step of ribozyme self-cleavage

Ribozymes of hepatitis delta virus have been proposed to use an active-site cytosine as an acid-base catalyst in the self-cleavage reaction. In this study, we have examined the role of cytosine in more detail with the antigenomic ribozyme. Evidence that proton transfer in the rate-determining step involved cytosine 76 (C76) was obtained from examining cleavage activity of the wild-type and imidazole buffer-rescued C76-deleted (C76Δ) ribozymes in D2O and H2O. In both reactions, a similar kinetic isotope effect and shift in the apparent pKa indicate that the buffer is functionally substituting for the side chain in proton transfer. Proton inventory of the wild-type reaction supported a mechanism of a single proton transfer at the transition state. This proton transfer step was further characterized by exogenous base rescue of a C76Δ mutant with cytosine and imidazole analogues. For the imidazole analogues that rescued activity, the apparent pKa of the rescue reaction, measured under kcat/KM conditions, correlated with the pKa of the base. From these data a Brønsted coefficient (β) of 0.51 was determined for the base-rescued reaction of C76Δ. This value is consistent with that expected for proton transfer in the transition state. Together, these data provide strong support for a mechanism where an RNA side chain participates directly in general acid or general base catalysis of the wild-type ribozyme to facilitate RNA cleavage.

How flowering plants discriminate between self and non-self pollen to prevent inbreeding.

Flowering plants have evolved various genetic mechanisms to circumvent the tendency for self-fertilization created by the close proximity of male and female reproductive organs in a bisexual flower. One such mechanism is gametophytic self-incompatibility, which allows the female reproductive organ, the pistil, to distinguish between self pollen and non-self pollen; self pollen is rejected, whereas non-self pollen is accepted for fertilization. The Solanaceae family has been used as a model to study the molecular and biochemical basis of self/non-self-recognition and self-rejection. Discrimination of self and non-self pollen by the pistil is controlled by a single polymorphic locus, the S locus. The protein products of S alleles in the pistil, S proteins, were initially identified based on their cosegregation with S alleles. S proteins have recently been shown to indeed control the ability of the pistil to recognize and reject self pollen. S proteins are also RNases, and the RNase activity has been shown to be essential for rejection of self pollen, suggesting that the biochemical mechanism of self-rejection involves the cytotoxic action of the RNase activity. S proteins contain various numbers of N-linked glycans, but the carbohydrate moiety has been shown not to be required for the function of S proteins, suggesting that the S allele specificity determinant of S proteins lies in the amino acid sequence. The male component in self-incompatibility interactions, the pollen S gene, has not yet been identified. The possible nature of the pollen S gene product and the possible mechanism by which allele-specific rejection of pollen is accomplished are discussed.

Preferential self-association of basic fibroblast growth factor is stabilized by heparin during receptor dimerization and activation.

Central to signaling by fibroblast growth factors (FGFs) is the oligomeric interaction of the growth factor and its high-affinity cell surface receptor, which is mediated by heparin-like polysaccharides. It has been proposed that the binding of heparin-like polysaccharides to FGF induces a conformational change in FGF, resulting in the formation of FGF dimers or oligomers, and this biologically active form is 'presented' to the FGF receptor for signal transduction. In this study, we show that monomeric basic FGF (FGF-2) preferentially self-associates and forms FGF-2 dimers and higher-order oligomers. As a consequence, FGF-2 monomers are oriented for binding to heparin-like polysaccharides. We also show that heparin-like polysaccharides can readily bind to self-associated FGF-2 without causing a conformational change in FGF-2 or disrupting the FGF-2 self-association, but that the bound polysaccharides only additionally stabilize the FGF-2 self-association. The preferential self-association corresponds to FGF-2 translations along two of the unit cell axes of the FGF-2 crystal structures. These two axes represent the two possible heparin binding directions, whereas the receptor binding sites are oriented along the third axis. Thus, we propose that preferential FGF-2 self-association, further stabilized by heparin, like "beads on a string," mediates FGF-2-induced receptor dimerization and activation. The observed FGF-2 self-association, modulated by heparin, not only provides a mechanism of growth factor activation but also represents a regulatory mechanism governing FGF-2 biological activity.

Antagonism of WT1 activity by protein self-association.

Germline loss-of-function mutations at the Wilms tumor (WT) suppressor locus WT1 are associated with a predisposition to WTs and mild genital system anomalies. In contrast, germ-line missense mutations within the WT1 gene encoding the DNA-binding domain often yield a more severe phenotype consisting of WT, sexual ambiguity, and renal nephropathy. In this report, we demonstrate that the products of mutant alleles that impair DNA recognition can antagonize WT1-mediated transcriptional repression. We demonstrate that WT1 can self-associate in vitro and in vivo and that the responsible domain maps to the amino-terminal region of the protein. Oligomers of full-length protein form less efficiently or produce less stable complexes than oligomers between truncated polypeptides and full-length protein. Our data suggest a molecular mechanism to explain how WT1 mutations may act in deregulating cellular proliferation and differentiation.