Biomedical subjects

Akira Chiba

Publications and source records attributed to Akira Chiba.

13 recordsLinked to original sources

PubMed2007

Nanomolar inhibition of type II dehydroquinase based on the enolate reaction mechanism.

We describe the rational design of a novel, highly potent inhibitor of type II dehydroquinase, the dicarboxylate 6. The incorporation of a carboxylate at the 3-position mimics the putative enolate intermediate in the reaction mechanism, and allows a potential electrostatic binding interaction with the arginine on the active site flap. This results in a 1000-fold increase in potency, making the dicarboxylate 6 the most potent inhibitor of type II dehydroquinase reported to date, with a high ligand efficiency of -0.68 kcal mol(-1) per nonhydrogen atom. The systematic dissection of 6 in compounds 7-12, all of which show a drop in potency, confirm the synergistic importance of the two carboxylates, the C3 and C4 hydroxyl groups, and the anhydroquinate ring structure for the potency of 6.

Alcohols↗

PubMed2006

The variable transmembrane domain of Drosophila N-cadherin regulates adhesive activity.

Drosophila N-cadherin (CadN) is an evolutionarily conserved classic cadherin which has a large, complex extracellular domain and a catenin-binding cytoplasmic domain. The CadN locus contains three modules of alternative exons (7a/b, 13a/b, and 18a/b) and undergoes alternative splicing to generate multiple isoforms. Using quantitative transcript analyses and green fluorescent protein-based cell sorting, we found that during development CadN alternative splicing is regulated in a temporal but not cell-type-specific fashion. In particular, exon 18b is predominantly expressed during early developmental stages, while exon 18a is prevalent at the late developmental and adult stages. All CadN isoforms share the same molecular architecture but have different sequences in their extracellular and transmembrane domains, suggesting functional diversity. In vitro quantitative cell aggregation assays revealed that all CadN isoforms mediate homophilic interactions, but the isoforms encoded by exon 18b have a higher adhesive activity than those by its alternative, 18a. Domain-swapping experiments further revealed that the different sequences in the transmembrane domains of isoforms are responsible for their differential adhesive activities. CadN alternative splicing might provide a novel mechanism to fine-tune its adhesive activity at different developmental stages or to restrict the use of high-affinity 18b-type isoforms at the adult stage.

Alternative Splicing↗

PubMed2005

Gliopodia extend the range of direct glia-neuron communication during the CNS development in Drosophila.

Midline glia are a source of cues for neuronal navigation and differentiation in the Drosophila CNS. Despite their importance, how glia and neurons communicate during the development is not fully understood. Here, we examined dynamic morphology of midline glia and assessed their direct cellular interactions with neurons within the embryonic CNS. Midline glia extend filopodia-like "gliopodia" from the onset of axogenesis through the near completion of embryonic neural development. The most abundant and stable within the commissures, gliopodia frequently contact neurites extending from the neuropil on either side of the midline. Misexpression of Rac1N17 in midline glia not only reduces the number of gliopodia but also shifts the position of neuropils towards the midline. Midline-secreted signaling protein Slit accumulates along the surface of gliopodia. Mutant analysis supports the idea that gliopodia contribute to its presentation on neuronal surfaces at both the commissures and neuropils. We propose that gliopodia extend the range of direct glia-neuron communication during CNS development.

Animals↗

PubMed2005

Drosophila N-cadherin functions in the first stage of the two-stage layer-selection process of R7 photoreceptor afferents.

Visual information received from the three types of photoreceptor neurons (R1-R6, R7 and R8) in the fly compound eyes converges to the external part of the medulla neuropil (M1-M6 layers) in a layer-specific fashion: R7 and R8 axons terminate at the M6 and M3 layers, respectively, whereas lamina neurons (L1-L5) relay R1-R6 to multiple medulla layers (M1-M5). Here, we show that during development, R7 and R8 neurons establish layer-specific projections in two separate stages: during the first stage, R7 and R8 axons sequentially target to the R7- and R8-temporary layers, respectively; and at the second stage, R7 and R8 growth cones progress synchronously to their destined layers. Using a set of mutations that delete different afferent subsets or alter R7 connectivity, we defined the mechanism of layer selection. We observed that R8, R7 and L1-L5 afferents target to their temporary layers independently, suggesting that afferent-target, but not afferent-afferent, interactions dictate the targeting specificity. N-cadherin is required in the first stage for R7 growth cones to reach and remain in the R7-temporary layer. The Ncad gene contains three pairs of alternatively spliced exons and encodes 12 isoforms. However, expressing a single Ncad isoform in Ncad mutant R7s is sufficient to rescue mistargeting phenotypes. Furthermore, Ncad isoforms mediate promiscuous heterophilic interactions in an in vitro cell-aggregation assay. We propose that Ncad isoforms do not form an adhesion code; rather, they provide permissive adhesion between R7 growth cones and their temporary targets.

Alternative Splicing↗

PubMed2005

A novel cysteine protease inhibitor with lectin activity from the epidermis of the Japanese eel Anguilla japonica.

A novel cysteine protease inhibitor (Eel-CPI-1) was isolated from the epidermis of the eel. Eel-CPI-1 was shown to bind strongly to both lactose- and carboxymethylated papain-affinity gels. Its molecular mass under reducing condition was determined to be 18 kDa by SDS-polyacrylamide gel electrophoresis but approximately 30.5 kDa under non-reducing-conditions. Eel-CPI-1 inhibited papain (K(i)=18 nM) and ficin (K(i)=120 nM) competitively. Combined with the data on amino acid and sequence analysis, Eel-CPI-1 is identical to the eel lectin, AJL-2. This is the first report describing a cysteine protease inhibitor with lectin activity.

Animals↗

PubMed2004

Dendritic guidance.

Like axons, dendrites need guidance for proper orientation and positioning within the brain. Guidance determines synaptic connectivity as well as the strength of transmission. Recent in vivo studies have demonstrated that several cell-surface receptors, previously known as axon guidance molecules, are also responsible for the directed outgrowth of dendrites. Collectively, these studies reveal that the function of guidance molecules in individual neurons and individual processes is diverse and likely to be specifically regulated. Here, these studies are reviewed and emerging issues and implications are discussed.

Animals↗

PubMed2003

Isolation of Rho GTPase effector pathways during axon development.

The Rho GTPases Rac1 and Cdc42 have been implicated in the regulation of axon outgrowth and guidance. However, the downstream effector pathways through which these GTPases exert their effects on axon development are not well characterized. Here, we report that axon outgrowth defects within specific subsets of motoneurons expressing constitutively active Drosophila Rac1 largely persist even with the addition of an effector-loop mutation to Rac1 that disrupts its ability to bind to p21-activated kinase (Pak) and other Cdc42/Rac1 interactive-binding (CRIB)-motif effector proteins. While hyperactivation of Pak itself does not lead to axon outgrowth defects as when Rac1 is constitutively activated, live analysis reveals that it can alter filopodial activity within specific subsets of neurons similar to constitutive activation of Cdc42. Moreover, we show that the axon guidance defects induced by constitutive activation of Cdc42 persist even in the absence of Pak activity. Our results suggest that (1) Rac1 controls axon outgrowth through downstream effector pathways distinct from Pak, (2) Cdc42 controls axon guidance through both Pak and other CRIB effectors, and (3) Pak's primary contribution to in vivo axon development is to regulate filopodial dynamics that influence growth cone guidance.

Animals↗

PubMed2003

Myopodia (postsynaptic filopodia) participate in synaptic target recognition.

Synaptic partner cells recognize one another by utilizing a variety of molecular cues. Prior to neuromuscular synapse formation, Drosophila embryonic muscles extend dynamic actin-based filopodia called "myopodia." In wild-type animals, myopodia are initially extended randomly from the muscle surface but become gradually restricted to the site of motoneuron innervation, a spatial redistribution we call "clustering." Previous experiments with prospero mutant embryos demonstrated that myopodia clustering does not occur in the absence of motoneuron outgrowth into the muscle field. However, whether myopodia clustering is due to a general signal from passing axons or is a result of the specific interactions between synaptic partners remained to be investigated. Here, we have examined the relationship of myopodia to the specific events of synaptic target recognition, the stable adhesion of synaptic partners. We manipulated the embryonic expression of alphaPS2 integrin and Toll, molecules known to affect synaptic development, to specifically alter synaptic targeting on identified muscles. Then, we used a vital single-cell labeling approach to visualize the behavior of myopodia in these animals. We demonstrate a strong positive correlation between myopodia activity and synaptic target recognition. The frequency of myopodia clustering is lowered in cases where synaptic targeting is disrupted. Myopodia clustering seems to result from the adherence of a subset of myopodia to the innervating growth cone while the rest are eliminated. The data suggest that postsynaptic cells play a dynamic role in the process of synaptic target recognition.

Amino Acids↗

PubMed2003

Robo and Frazzled/DCC mediate dendritic guidance at the CNS midline.

Neuronal connectivity is established by the axo-dendritic polarity, correct guidance and targeting of neurons. Unlike for axons, the mechanisms responsible for directed outgrowth of dendrites are not well understood. Using single-cell labeling, we describe specific guidance defects in dendrites of identified neurons in frazzled, robo, netrin and commissureless mutant embryos of Drosophila melanogaster. We found that the cell-surface molecules Frazzled and Robo work as guidance molecules not only for axons but also for dendrites as they navigate within the CNS. Furthermore, we report that each neuron showed a cell-autonomous and independent use of guidance molecules.

Animals↗

PubMed2003

Exclusion of ribulose-1,5-bisphosphate carboxylase/oxygenase from chloroplasts by specific bodies in naturally senescing leaves of wheat.

Immunocytochemical electron-microscopic observation indicated that ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco, EC 4.1.1.39) and/or its degradation products are localized in small spherical bodies having a diameter of 0.4-1.2 micro m in naturally senescing leaves of wheat (Triticum aestivum L.). These Rubisco-containing bodies (RCBs) were found in the cytoplasm and in the vacuole. RCBs contained another stromal protein, chloroplastic glutamine synthetase, but not thylakoid proteins. Ultrastructural analysis suggested that RCBs had double membranes, which seemed to be derived from the chloroplast envelope, and that RCBs were further surrounded by the other membrane structures in the cytoplasm. The appearance of RCBs was the most remarkable when the amount of Rubisco started to decrease at the early phase of leaf senescence. These results suggest that RCBs might be involved in the degradation process of Rubisco outside of chloroplasts during leaf senescence.

Hydrolysis↗

PubMed2002

Ontogenetic changes in neuropeptide Y-immunoreactive cerebrospinal fluid-contacting neurons in the hypothalamus of the cloudy dogfish, Scyliorhinus torazame (Elasmobranchii).

Ontogenetic changes in neuropeptide Y-immunoreactive (NPY-ir) cerebrospinal fluid (CSF)-contacting neurons in the dogfish hypothalamus were studied immunohistochemically. NPY-ir CSF-contacting neurons first appeared in the median infundibular floor of the embryo at the 34 mm stage. At the 40 mm stage, similar neurons were found also in the saccus vasculosus (SV). The number of these neurons increased during the 54-80 mm stages, and the cells in the infundibular floor extended their basal processes to the neuropil of the median eminence, whereas the cells in the SV sent their axonal fibers to the tractus sacci vasculosi. After hatching, NPY immunoreactivity in the ventral hypothalamus became less dense, and the labeled CSF-contacting neurons tended to be confined to the nucleus lateralis tuberis, similarly as in the adults. The occurrence of NPY-ir CSF-contacting neurons in the SV was transient during the embryonic periods.

Animals↗

PubMed2002

Growth cone pathfinding and filopodial dynamics are mediated separately by Cdc42 activation.

Although evidence exists that activation of the Rho family GTPase Cdc42 affects axonal development, its specific roles within a growth cone are not well delineated. To evaluate the model that Cdc42 activation regulates growth cone navigation by promoting filopodial activity, we adopted a live analysis strategy that uses transgenic Drosophila lines in which neurons coexpressed constitutively active Cdc42 (Cdc42(V12)) and membrane-targeted green fluorescent protein. We found that growth cones that displayed pathfinding defects exhibited little change in their filopodial activity, whereas others without pathfinding defects exhibited an similar50% increase in their filopodial activity. Moreover, effector loop mutations that were added to the constitutively active Cdc42 (Cdc42(V12C40) and Cdc42(V12A37)) exerted little influence over filopodial activity caused by Cdc42 activation but suppressed the pathfinding defects of the growth cones. Together, these data suggest that Cdc42 controls filopodial activity in axonal growth cones independently of its effects on their pathfinding.

Amino Acid Substitution↗

PubMedDate not supplied

Molecular mechanisms for Drosophila neuronetwork formation.

Understanding the establishment of functional neuronetworks is one of the frontiers of developmental neurobiology. The use of axons and dendrites from Drosophila nervous system as a model allows the identification of molecular mechanisms which give neurons the ability to guide their processes en route to connect precisely with their partners. By focusing on selected Drosophila model systems, we discuss the recent advances in our understanding of the molecular mechanisms regulating guidance, branching and targeting of axons and dendrites required for the establishment of a functional neuronetwork.

Animals↗