Three distinct modes of exocytosis revealed by amperometry in neuroendocrine cells

Neurotransmission requires Ca²⁺-dependent release of secretory products through fusion pores that open and reclose (partial membrane distention) or open irreversibly (complete membrane distention). It has been challenging to distinguish between these release modes; however, in the work presented here, we were able to deduce different modes of depolarization-evoked exocytosis in neuroendocrine chromaffin and PC12 cells solely by analyzing amperometric recordings. After we determined the quantal size (Q), event half-width (t₅₀), event amplitude (I_peak), and event decay time constant (τ_decay), we fitted scatter plots of log-transformed data with a mixture of one- and two-dimensional Gaussian distributions. Our analysis revealed three distinct and differently shaped clusters of secretory events, likely corresponding to different modes of exocytosis. Complete membrane distention, through fusion pores of widely varying conductances, accounted for 70% of the total amount of released catecholamine. Two different kinds of partial membrane distention (kiss-and-run and kiss-and-stay exocytosis), characterized by mode-specific fusion pores with unitary conductances, accounted for 20% and 10%, respectively. These results show that our novel one- and two-dimensional analysis of amperometric data reveals new release properties and enables one to distinguish at least three different modes of exocytosis solely by analyzing amperometric recordings.     

EEG data acquisition and preprocessing by microcomputer satellite system

Recent development in computer technology allows already medium scale EEG data processing to be performed within the clinical neurophysiology department, if a fast minicomputer with adequate mass storage and graphical output facilities is used. Data acquisition, however, should be delegated to a microcomputer which also should take over as much preprocessing as possible. A system is presented, where one or several microcomputer-based satellite units perform analog-digital conversion, Fourier transformation (FFT), calculation of power spectra and crossproducts, as well as event related averaging or other preprocessing procedures. The units are connected to a fast central minicomputer, where a supervisor program loads the microprocessors with their programs, supervises their activity, receives preprocessed data and activates appropriate postprocessing programs to produce the final results.     

Three modes of asexual reproduction in the moss Octoblepharum albidum

Abstract

Populations of Octoblepharum albidum growing in very moist and shaded habitats were found to have three kinds of asexual reproduction. First, the production of foliar gemmae along the upper half of leaf margins. Second, the production of protonema gemmae on protonemata originated from foliar gemmae. Third, the production of buds and new shoots at leaf tips. The formation of new shoots is further repeated at the leaf tips of the new shoots, giving the plant the character of a 'walking moss'.     

Personal identification by eyes

Identification of persons through the eyes is in the field of biometrical science. Many security systems are based on biometric methods of personal identification, to determine whether a person is presenting itself truly. The human eye contains an extremely large number of individual characteristics that make it particularly suitable for the process of identifying a person. Today, the eye is considered to be one of the most reliable body parts for human identification. Systems using iris recognition are among the most secure biometric systems.     

A preprocessing procedure for haplotype inference by pure parsimony

Haplotype data are especially important in the study of complex diseases since it contains more information than genotype data. However, obtaining haplotype data is technically difficult and costly. Computational methods have proved to be an effective way of inferring haplotype data from genotype data. One of these methods, the haplotype inference by pure parsimony approach (HIPP), casts the problem as an optimization problem and as such has been proved to be NP-hard. We have designed and developed a new preprocessing procedure for this problem. Our proposed algorithm works with groups of haplotypes rather than individual haplotypes. It iterates searching and deleting haplotypes that are not helpful in order to find the optimal solution. This preprocess can be coupled with any of the current solvers for the HIPP that need to preprocess the genotype data. In order to test it, we have used two state-of-the-art solvers, RTIP and GAHAP, and simulated and real HapMap data. Due to the computational time and memory reduction caused by our preprocess, problem instances that were previously unaffordable can be now efficiently solved.     

Three modes of age-related changes of hydrolytic enzymes in murine brain

We followed the time course of the activities of various hydrolytic enzymes in murine brain for 30 months to investigate their roles in the aging process. Although most of the 15 enzymatic activities tested tended to rise with the increase in age, by analyzing them by multivariate analysis their movements could be divided into three types, each having a particular mode of nonlinear regression: exponential, logarithmic, or parabolic. Several enzymatic activities, including those of angiotensin-converting enzyme, esterase, trypsin-like enzyme, post-proline-cleaving enzyme, and Gly-Pro-aminopeptidase, showed rhythmic oscillations with about one cycle per 3 months throughout the period of 30 months. The behavior of these enzymes probably represents some particular aspect of metabolism relatively independent from the aging process. These findings may provide a fundamental clue in association with the process of maturation and aging in the brain.     

Monitoring spatiotemporal biogenesis of macromolecular assemblies by pulse-chase epitope labeling

Many cellular proteins perform their roles within macromolecular assemblies. Hence, an understanding of how these multiprotein complexes form is?a fundamental question in cell biology. We developed a translation-controlled pulse-chase system that allows time-resolved isolation of newly forming multiprotein complexes in chemical quantities suitable for biochemical and cell biological analysis. The "pulse" is triggered by an unnatural amino acid, which induces immediate translation of an amber stop codon repressed mRNA encoding the protein of interest with a built-in tag for detection and purification. The "chase" is elicited by stopping translation of this bait via a riboswitch in the respective mRNA. Over the course of validating our method, we discovered a distinct time-resolved assembly step during NPC biogenesis and could directly monitor the spatiotemporal maturation of preribosomes via immunofluorescence detection and purification of a pulse-labeled ribosomal protein. Thus, we provide an innovative strategy to study dynamic protein assembly within cellular networks.     

Regulation of nuclear PKA revealed by spatiotemporal manipulation of cyclic AMP

Understanding how specific cyclic AMP (cAMP) signals are organized and relayed to their effectors in different compartments of the cell to achieve functional specificity requires molecular tools that allow precise manipulation of cAMP in these compartments. Here we characterize a new method using bicarbonate-activatable and genetically targetable soluble adenylyl cyclase to control the location, kinetics and magnitude of the cAMP signal. Using this live-cell cAMP manipulation in conjunction with fluorescence imaging and mechanistic modeling, we uncovered the activation of a resident pool of protein kinase A (PKA) holoenzyme in the nuclei of HEK-293 cells, modifying the existing dogma of cAMP-PKA signaling in the nucleus. Furthermore, we show that phosphodiesterases and A-kinase anchoring proteins (AKAPs) are critical in shaping nuclear PKA responses. Collectively, our data suggest a new model in which AKAP-localized phosphodiesterases tune an activation threshold for nuclear PKA holoenzyme, thereby converting spatially distinct second messenger signals to temporally controlled nuclear kinase activity.     

Quality control and preprocessing of metagenomic datasets

SUMMARY: Here, we present PRINSEQ for easy and rapid quality control and data preprocessing of genomic and metagenomic datasets. Summary statistics of FASTA (and QUAL) or FASTQ files are generated in tabular and graphical form and sequences can be filtered, reformatted and trimmed by a variety of options to improve downstream analysis. Availability and Implementation: This open-source application was implemented in Perl and can be used as a stand alone version or accessed online through a user-friendly web interface. The source code, user help and additional information are available at http://prinseq.sourceforge.net/.     

Genetic network inference: the effects of preprocessing

Clustering of gene expression data and gene network inference from such data has been a major research topic in recent years. In clustering, pairwise measurements are performed when calculating the distance matrix upon which the clustering is based. Pairwise measurements can also be used for gene network inference, by deriving potential interactions above a certain correlation or distance threshold. Our experiments show how interaction networks derived by this simple approach exhibit low-but significant-sensitivity and specificity. We also explore the effects that normalization and prefiltering have on the results of methods for identifying interactions from expression data. Before derivation of interactions or clustering, preprocessing is often performed by applying normalization to rescale the expression profiles and prefiltering where genes that do not appear to contribute to regulation are removed. In this paper, different ways of normalizing in combination with different distance measurements are tested on both unfiltered and prefiltered data, different prefiltering criteria are considered.     

Two modes of ligand recognition by TLRs

Toll-like receptors (TLRs) are membrane-bound sensors that detect and respond to microbial infection. Two studies in Cell, one in this issue, reveal how TLRs recognize their ligands. Kim et al. (2007) recently reported the structure of TLR4 bound to the accessory protein MD-2 and its antagonist, the drug eritoran. In this issue, Jin et al. (2007) describe the crystal structure of a complex between TLR1, TLR2, and a lipopeptide ligand.     

Stabilization of ballooning modes by nonparaxial cells

An analysis is made of the effect of high-curvature stabilizing nonparaxial elements (cells) on the MHD plasma stability in open confinement systems and in confinement systems with closed magnetic field lines. It is shown that the population of particles trapped in such cells has a stabilizing effect not only on convective (flute) modes but also on ballooning modes, which govern the maximum possible β value. In the kinetic approach, which distinguishes between the effects of trapped and passing particles, the maximum possible β values consistent with stability can be much higher than those predicted by the MHD model.     

Limb,tooth, beak: Three modes of development and evolutionary innovation of form

The standard model of evolutionary change of form, deriving from Darwin’s theory via the Modern Synthesis, assumes a gradualistic reshaping of anatomical structures, with major changes only occurring by many cycles of natural selection for marginal adaptive advantage. This model, with its assertion that a single mechanism underlies both micro- and macroevolutionary change, contains an implicit notion of development which is only applicable in some cases. Here we compare the embryological processes that shape the vertebrate limb bud, the mammalian tooth and the avian beak. The implied notion of development in the standard evolutionary picture is met only in the case of the vertebrate limb, a single-primordium organ with morphostatic shaping, in which cells rearrange in response to signalling centres which are essentially unchanged by cell movement. In the case of the tooth, a single-primordium organ with morphodynamic shaping in which the strengths and relationships between signalling centres is influenced by the cell and tissue movements they induce, and the beak, in which the final form is influenced by the collision and rearrangement of multiple tissue primordia, abrupt appearance of qualitatively different forms (i.e. morphological novelties) can occur with small changes in system parameters induced by a genetic change, or by an environmental factor whose effects can be subsequently canalized genetically. Bringing developmental mechanisms and, specifically, the material properties of tissues as excitable media into the evolutionary picture, demonstrates that gradualistic change for incremental adaptive advantage is only one of the possible modes of morphological evolution.     

Cognitive association formation in human memory revealed by spatiotemporal brain imaging

Cognitive theory posits association by juxtaposition or by fusion. We employed the measurement of event-related brain potentials (ERPs) to a concept fusion task in order to explore memory encoding of these two types of associations between word pairs, followed by a memory test for original pair order. Encoding processes were isolated by subtracting fusion task ERPs corresponding to pairs later retrieved quickly from ERPs corresponding to pairs later retrieved slowly, separately for pairs fused successfully and unsuccessfully (i.e., juxtaposed). Analyses revealed that the encoding of these two types of associations yields different ERP voltage polarities, scalp topographies, and brain sources extending over the entire time course of processing.