16 results on '"Kamasawa, Naomi"'
Search Results
2. Cortical response selectivity derives from strength in numbers of synapses
- Author
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Scholl, Benjamin, Thomas, Connon I., Ryan, Melissa A., Kamasawa, Naomi, and Fitzpatrick, David
- Abstract
Single neocortical neurons are driven by populations of excitatory inputs, which form the basis of neuronal selectivity to features of sensory input. Excitatory connections are thought to mature during development through activity-dependent Hebbian plasticity1, whereby similarity between presynaptic and postsynaptic activity selectively strengthens some synapses and weakens others2. Evidence in support of this process includes measurements of synaptic ultrastructure and in vitro and in vivo physiology and imaging studies3–8. These corroborating lines of evidence lead to the prediction that a small number of strong synaptic inputs drive neuronal selectivity, whereas weak synaptic inputs are less correlated with the somatic output and modulate activity overall6,7. Supporting evidence from cortical circuits, however, has been limited to measurements of neighbouring, connected cell pairs, raising the question of whether this prediction holds for a broad range of synapses converging onto cortical neurons. Here we measure the strengths of functionally characterized excitatory inputs contacting single pyramidal neurons in ferret primary visual cortex (V1) by combining in vivo two-photon synaptic imaging and post hoc electron microscopy. Using electron microscopy reconstruction of individual synapses as a metric of strength, we find no evidence that strong synapses have a predominant role in the selectivity of cortical neuron responses to visual stimuli. Instead, selectivity appears to arise from the total number of synapses activated by different stimuli. Moreover, spatial clustering of co-active inputs appears to be reserved for weaker synapses, enhancing the contribution of weak synapses to somatic responses. Our results challenge the role of Hebbian mechanisms in shaping neuronal selectivity in cortical circuits, and suggest that selectivity reflects the co-activation of large populations of presynaptic neurons with similar properties and a mixture of strengths.
- Published
- 2021
- Full Text
- View/download PDF
3. LRIT1 Modulates Adaptive Changes in Synaptic Communication of Cone Photoreceptors
- Author
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Sarria, Ignacio, Cao, Yan, Wang, Yuchen, Ingram, Norianne T., Orlandi, Cesare, Kamasawa, Naomi, Kolesnikov, Alexander V., Pahlberg, Johan, Kefalov, Vladimir J., Sampath, Alapakkam P., and Martemyanov, Kirill A.
- Abstract
Cone photoreceptors scale dynamically the sensitivity of responses to maintain responsiveness across wide range of changes in luminance. Synaptic changes contribute to this adaptation, but how this process is coordinated at the molecular level is poorly understood. Here, we report that a cell adhesion-like molecule, LRIT1, is enriched selectively at cone photoreceptor synapses where it engages in a trans-synaptic interaction with mGluR6, the principal receptor in postsynaptic ON-bipolar cells. The levels of LRIT1 are regulated by the neurotransmitter release apparatus that controls photoreceptor output. Knockout of LRIT1 in mice increases the sensitivity of cone synaptic signaling while impairing its ability to adapt to background light without overtly influencing the morphology or molecular composition of photoreceptor synapses. Accordingly, mice lacking LRIT1 show visual deficits under conditions requiring temporally challenging discrimination of visual signals in steady background light. These observations reveal molecular mechanisms involved in scaling synaptic communication in the retina.
- Published
- 2018
- Full Text
- View/download PDF
4. Retrograde Synaptic Signaling Mediated by K+Efflux through Postsynaptic NMDA Receptors
- Author
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Shih, Pei-Yu, Savtchenko, Leonid P., Kamasawa, Naomi, Dembitskaya, Yulia, McHugh, Thomas J., Rusakov, Dmitri A., Shigemoto, Ryuichi, and Semyanov, Alexey
- Abstract
Synaptic NMDA receptors (NMDARs) carry inward Ca2+current responsible for postsynaptic signaling and plasticity in dendritic spines. Whether the concurrent K+efflux through the same receptors into the synaptic cleft has a physiological role is not known. Here, we report that NMDAR-dependent K+efflux can provide a retrograde signal in the synapse. In hippocampal CA3-CA1 synapses, the bulk of astrocytic K+current triggered by synaptic activity reflected K+efflux through local postsynaptic NMDARs. The local extracellular K+rise produced by activation of postsynaptic NMDARs boosted action potential-evoked presynaptic Ca2+transients and neurotransmitter release from Schaffer collaterals. Our findings indicate that postsynaptic NMDAR-mediated K+efflux contributes to use-dependent synaptic facilitation, thus revealing a fundamental form of retrograde synaptic signaling.
- Published
- 2013
- Full Text
- View/download PDF
5. Cell Surface Engineering Enables Surfaceome Profiling
- Author
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Vilen, Zak, Reeves, Abigail E., O’Leary, Timothy R., Joeh, Eugene, Kamasawa, Naomi, and Huang, Mia L.
- Abstract
Cell surface proteins (CSPs) are vital molecular mediators for cells and their extracellular environment. Thus, understanding which CSPs are displayed on cells, especially in different cell states, remains an important endeavor in cell biology. Here, we describe the integration of cell surface engineering with radical-mediated protein biotinylation to profile CSPs. This method relies on the prefunctionalization of cells with cholesterol lipid groups, followed by sortase-catalyzed conjugation with an APEX2 ascorbate peroxidase enzyme. In the presence of biotin-phenol and H2O2, APEX2 catalyzes the formation of highly reactive biotinyl radicals that covalently tag electron-rich residues within CSPs for subsequent streptavidin-based enrichment and analysis by quantitative mass spectrometry. While APEX2 is traditionally used to capture proximity-based interactomes, we envisioned using it in a “baitless” manner on cell surfaces to capture CSPs. We evaluate this strategy in light of another CSP labeling method that relies on the presence of cell surface sialic acid. Using the APEX2 strategy, we describe the CSPs found in three mammalian cell lines and compare CSPs in adherent versus three-dimensional pancreatic adenocarcinoma cells.
- Published
- 2022
- Full Text
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6. <TOGGLE>In situ</TOGGLE> localization of β-glucans in the cell wall of <TOGGLE>Schizosaccharomyces pombe</TOGGLE>
- Author
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Humbel, Bruno M., Konomi, Mami, Takagi, Tomoko, Kamasawa, Naomi, Ishijima, Sanae A., and Osumi, Masako
- Abstract
The chemical composition of the cell wall of Sz. pombe is known as β-1,3-glucan, β-1,6-glucan, α-1,3-glucan and α-galactomannan; however, the three-dimensional interactions of those macromolecules have not yet been clarified. Transmission electron microscopy reveals a three-layered structure: the outer layer is electron-dense, the adjacent layer is less dense, and the third layer bordering the cell membrane is dense. In intact cells of Sz. pombe, the high-resolution scanning electron microscope reveals a surface completely filled with α-galactomannan particles. To better understand the organization of the cell wall and to complement our previous studies, we set out to locate the three different types of β-glucan by immuno-electron microscopy. Our results suggest that the less dense layer of the cell wall contains mainly β-1,6-branched β-1,3-glucan. Occasionally a line of gold particles can be seen, labelling fine filaments radiating from the cell membrane to the α-galactomannan layer, suggesting that some of the radial filaments contain β-1,6-branched β-1,3-glucan. β-1,6-glucan is preferentially located underneath the α-galactomannan layer. Linear β-1,3-glucan is exclusively located in the primary septum of dividing cells. β-1,6-glucan only labels the secondary septum and does not co-localize with linear β-1,3-glucan, while β-1,6-branched β-1,3-glucan is present in both septa. Linear β-1,3-glucan is present from early stages of septum formation and persists until the septum is completely formed; then just before cell division the label disappears. From these results we suggest that linear β-1,3-glucan is involved in septum formation and perhaps the separation of the two daughter cells. In addition, we frequently found β-1,6-glucan label on the Golgi apparatus, on small vesicles and underneath the cell membrane. These results give fresh evidence for the hypothesis that β-1,6-glucan is synthesized in the endoplasmic reticulumGolgi system and exported to the cell membrane. Copyright © 2001 John Wiley & Sons, Ltd.
- Published
- 2001
7. In situlocalization of β‐glucans in the cell wall of Schizosaccharomyces pombe
- Author
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Humbel, Bruno M., Konomi, Mami, Takagi, Tomoko, Kamasawa, Naomi, Ishijima, Sanae A., and Osumi, Masako
- Abstract
The chemical composition of the cell wall of Sz. pombeis known as β‐1,3‐glucan, β‐1,6‐glucan, α‐1,3‐glucan and α‐galactomannan; however, the three‐dimensional interactions of those macromolecules have not yet been clarified. Transmission electron microscopy reveals a three‐layered structure: the outer layer is electron‐dense, the adjacent layer is less dense, and the third layer bordering the cell membrane is dense. In intact cells of Sz. pombe, the high‐resolution scanning electron microscope reveals a surface completely filled with α‐galactomannan particles. To better understand the organization of the cell wall and to complement our previous studies, we set out to locate the three different types of β‐glucan by immuno‐electron microscopy. Our results suggest that the less dense layer of the cell wall contains mainly β‐1,6‐branched β‐1,3‐glucan. Occasionally a line of gold particles can be seen, labelling fine filaments radiating from the cell membrane to the α‐galactomannan layer, suggesting that some of the radial filaments contain β‐1,6‐branched β‐1,3‐glucan. β‐1,6‐glucan is preferentially located underneath the α‐galactomannan layer. Linear β‐1,3‐glucan is exclusively located in the primary septum of dividing cells. β‐1,6‐glucan only labels the secondary septum and does not co‐localize with linear β‐1,3‐glucan, while β‐1,6‐branched β‐1,3‐glucan is present in both septa. Linear β‐1,3‐glucan is present from early stages of septum formation and persists until the septum is completely formed; then just before cell division the label disappears. From these results we suggest that linear β‐1,3‐glucan is involved in septum formation and perhaps the separation of the two daughter cells. In addition, we frequently found β‐1,6‐glucan label on the Golgi apparatus, on small vesicles and underneath the cell membrane. These results give fresh evidence for the hypothesis that β‐1,6‐glucan is synthesized in the endoplasmic reticulum–Golgi system and exported to the cell membrane. Copyright © 2001 John Wiley & Sons, Ltd.
- Published
- 2001
- Full Text
- View/download PDF
8. Cytochemical evaluation of localization and secretion of a heterologous enzyme displayed on yeast cell surface
- Author
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Shibasaki, Yumi, Kamasawa, Naomi, Shibasaki, Seiji, Zou, Wen, Murai, Toshiyuki, Ueda, Mitsuyoshi, Tanaka, Atsuo, and Osumi, Masako
- Abstract
A starch‐utilizing Saccharomyces cerevisiaestrain was constructed by cell surface engineering. Distribution of the heterologous glucoamylase–α‐agglutinin fusion protein on the yeast cell was analyzed by indirect fluorescence microscopy using an anti‐glucoamylase antibody. Most of the intense fluorescence was first localized in the small bud, then observed on the entire cell wall of the daughter and mother cells. Fluorescence also accumulated at the neck region. These observations suggest that the display of the heterologous protein on the cell surface is carried with other cell wall components to the areas in which the cell wall is newly synthesized; the distribution is controlled by the cell cycle. Then, the heterologous protein‐encoding gene was expressed in a sec1mutant, in which secretory vesicles accumulate under restrictive temperature, and the produced protein was detected by immunoelectron microscopy. Most of the gold particles that reacted with the fusion protein were not localized in vesicles but in expanding endoplasmic reticulum. This phenomenon may be due to overproduction of the heterologous protein which was designed to be displayed on the cell wall. Artificial production of heterologous protein may have caused a relative shortage of glycosyl phosphatidylinositol anchors.
- Published
- 2000
- Full Text
- View/download PDF
9. Molecular Breeding of Polysaccharide-Utilizing Yeast Cells by Cell Surface Engineering
- Author
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UEDA, MITSUYOSHI, MURAI, TOSHIYUKI, SHIBASAKI, YUMI, KAMASAWA, NAOMI, OSUMI, MASAKO, and TANAKA, ATSUO
- Published
- 1998
- Full Text
- View/download PDF
10. Author Correction: Cortical response selectivity derives from strength in numbers of synapses
- Author
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Scholl, Benjamin, Thomas, Connon I., Ryan, Melissa A., Kamasawa, Naomi, and Fitzpatrick, David
- Abstract
A Correction to this paper has been published: https://doi.org/10.1038/s41586-020-03158-8.
- Published
- 2021
- Full Text
- View/download PDF
11. Computational modeling predicts ephemeral acidic microdomains in the glutamatergic synaptic cleft
- Author
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Feghhi, Touhid, Hernandez, Roberto X., Stawarski, Michal, Thomas, Connon I., Kamasawa, Naomi, Lau, A.W.C., and Macleod, Gregory T.
- Abstract
At chemical synapses, synaptic vesicles release their acidic contents into the cleft, leading to the expectation that the cleft should acidify. However, fluorescent pH probes targeted to the cleft of conventional glutamatergic synapses in both fruit flies and mice reveal cleft alkalinization rather than acidification. Here, using a reaction-diffusion scheme, we modeled pH dynamics at the Drosophilaneuromuscular junction as glutamate, ATP, and protons (H+) were released into the cleft. The model incorporates bicarbonate and phosphate buffering systems as well as plasma membrane calcium-ATPase activity and predicts substantial cleft acidification but only for fractions of a millisecond after neurotransmitter release. Thereafter, the cleft rapidly alkalinizes and remains alkaline for over 100 ms because the plasma membrane calcium-ATPase removes H+from the cleft in exchange for calcium ions from adjacent pre- and postsynaptic compartments, thus recapitulating the empirical data. The extent of synaptic vesicle loading and time course of exocytosis have little influence on the magnitude of acidification. Phosphate but not bicarbonate buffering is effective at suppressing the magnitude and time course of the acid spike, whereas both buffering systems are effective at suppressing cleft alkalinization. The small volume of the cleft levies a powerful influence on the magnitude of alkalinization and its time course. Structural features that open the cleft to adjacent spaces appear to be essential for alleviating the extent of pH transients accompanying neurotransmission.
- Published
- 2021
- Full Text
- View/download PDF
12. CAST/ELKS Proteins Control Voltage-Gated Ca2+Channel Density and Synaptic Release Probability at a Mammalian Central Synapse
- Author
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Dong, Wei, Radulovic, Tamara, Goral, R. Oliver, Thomas, Connon, Suarez Montesinos, Monica, Guerrero-Given, Debbie, Hagiwara, Akari, Putzke, Travis, Hida, Yamato, Abe, Manabu, Sakimura, Kenji, Kamasawa, Naomi, Ohtsuka, Toshihisa, and Young, Samuel M.
- Abstract
In the presynaptic terminal, the magnitude and location of Ca2+entry through voltage-gated Ca2+channels (VGCCs) regulate the efficacy of neurotransmitter release. However, how presynaptic active zone proteins control mammalian VGCC levels and organization is unclear. To address this, we deleted the CAST/ELKS protein family at the calyx of Held, a CaV2.1 channel-exclusive presynaptic terminal. We found that loss of CAST/ELKS reduces the CaV2.1 current density with concomitant reductions in CaV2.1 channel numbers and clusters. Surprisingly, deletion of CAST/ELKS increases release probability while decreasing the readily releasable pool, with no change in active zone ultrastructure. In addition, Ca2+channel coupling is unchanged, but spontaneous release rates are elevated. Thus, our data identify distinct roles for CAST/ELKS as positive regulators of CaV2.1 channel density and suggest that they regulate release probability through a post-priming step that controls synaptic vesicle fusogenicity.
- Published
- 2018
- Full Text
- View/download PDF
13. Cytochemical evaluation of localization and secretion of a heterologous enzyme displayed on yeast cell surface
- Author
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Shibasaki, Yumi, Kamasawa, Naomi, Shibasaki, Seiji, Zou, Wen, Murai, Toshiyuki, Ueda, Mitsuyoshi, Tanaka, Atsuo, and Osumi, Masako
- Abstract
A starch-utilizing Saccharomyces cerevisiae strain was constructed by cell surface engineering. Distribution of the heterologous glucoamylase–α-agglutinin fusion protein on the yeast cell was analyzed by indirect fluorescence microscopy using an anti-glucoamylase antibody. Most of the intense fluorescence was first localized in the small bud, then observed on the entire cell wall of the daughter and mother cells. Fluorescence also accumulated at the neck region. These observations suggest that the display of the heterologous protein on the cell surface is carried with other cell wall components to the areas in which the cell wall is newly synthesized; the distribution is controlled by the cell cycle. Then, the heterologous protein-encoding gene was expressed in a sec1 mutant, in which secretory vesicles accumulate under restrictive temperature, and the produced protein was detected by immunoelectron microscopy. Most of the gold particles that reacted with the fusion protein were not localized in vesicles but in expanding endoplasmic reticulum. This phenomenon may be due to overproduction of the heterologous protein which was designed to be displayed on the cell wall. Artificial production of heterologous protein may have caused a relative shortage of glycosyl phosphatidylinositol anchors.
- Published
- 2000
- Full Text
- View/download PDF
14. Three-dimensional analysis of protein aggregate body in Saccharomyces cerevisiae cells
- Author
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Kamasawa, Naomi, Yoshida, Tomoko, Ueda, Mitsuyoshi, Tanaka, Atsuo, and Osumi, Masako
- Abstract
Modified genes of peroxisomal isocitrate lyase of Candida tropicalis (CT-ICL) were constructed and expressed in Saccharomyces cerevisiae cells. We observed subcellular localization of expressed products of the mutant CT-ICL genes by immunoelectron microscopy. An unknown structure termed a protein aggregate body (PAB) storing the expressed product was observed in cytoplasm in various mutants (Kamasawa et al. (1996) J. Electron Microsc. 45: 491–497). We chose two typical cells harbouring the mutant ICL genes Δ550 and Δ237–339 to analyse the ultrastructure and three-dimensional (3D) structure of PABs. The PABs had a homogeneous matth with a wavy periphery in the cell image using a high-pressure freezing fixation method. Although PAB5 could not be separated from the cytoplasm or mitochondria under a confocal fluorescence microscope, 3D reconstruction of serial electron micrographs clearly showed the PAB was an independent structure of varying size and had the shape of an incomplete sphere. A cell was sometimes observed to have multiple PABs.
- Published
- 1999
- Full Text
- View/download PDF
15. Subcellular Destination of Mutant Peroxisomal Isocitrate Lyase Polypeptides of Candida tropicalis in Saccharomyces cerevisiae
- Author
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Kamasawa, Naomi, Yoshida, Tomoko, Kasahara, Mari, Kamada, Yasushi, Zou, Wen, Ueda, Mitsuyoshi, Tanaka, Atsuo, and Osumi, Masako
- Abstract
The peroxisomal isocitrate lyase (ICL) of an n-alkane-utilizable yeast, Candida tropicalis, could be transported into the peroxisomes of Saccharomyces cerevisiae MT8-1 cells grown in an oleic acid medium, even if strains were different. To identify the important regions to direct C. tropicalis ICL (CT-ICL) to its proper subcellular location in S. cerevisiae, we observed the subcellular localization of the expressed products of mutant CT-ICL genes in S. cerevisiae by immunoelectron microscopy. Under the control of the UPR-ICL promoter, which enhanced gene expression by proliferation of the peroxisomes, the following mutant CT-ICL genes were constructed and expressed: (i) truncated or mutant genes in the C-terminal region which apparently contained peroxisomal targeting signals, Δ550, Δ549–550, Δ548–550, Δ548–550 + SKL and Δ340–550; and (ii) four internal deletion mutant genes, Δ41–115, Δ41–239, Δ41–325 and Δ237–339. The results showed that three C-terminal amino acid residues were not essential for targeting the peroxisomes in this enzyme. In cells harboring the deleted internal regions of mutant ICL genes, the expressed polypeptides did not target the peroxisomes and the polypeptides, except for Δ41–115, were mistargeted to mitochondria. We observed an unknown structure which we called a “protein aggregate body (PAB),” in which various mutant CT-ICL polypeptides aggregated.
- Published
- 1996
- Full Text
- View/download PDF
16. Morphological Changes of Candida albicans Induced by BMY-28864, a Highly Water-soluble Pradimicin Derivative
- Author
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Numata, Keiichi, Ueki, Tomokazu, Naito, Nobuko, Yamada, Naoko, Kamasawa, Naomi, Oki, Toshikazu, and Osumi, Masako
- Abstract
The time course of BMY-28864-dependent morphological changes in Candida albicans A9540 was studied by electron microscopy. Scanning electron microscopy revealed that BMY-28864 often induced cell surface deformations such as abnormal swelling and bulging around budding sites and bud scars. The cell membrane damage was visualized by freeze-fracturing technique as deep pit-like invaginations. Transmission electron microscopy with thin-sectioned specimens also demonstrated that BMY-28864 induced cell membrane invaginations together with cell membrane detachment from the cell wall, nuclear membrane fragmentation and mitochondrial aberration. Statistical sequence analysis of the prominent BMY-28864-dependent morphological changes of the yeast cells led to the conclusion that BMY-28864 first attacked the cell membrane and then caused disintegration of other intracytoplasmic organelles, resulting in the lethal effect.
- Published
- 1993
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