Saitama University

9 ARTICLES PUBLISHED IN JoVE

Neuroscience

Morphological Analysis of Drosophila Larval Peripheral Sensory Neuron Dendrites and Axons Using Genetic Mosaics

¹Disease Mechanism Research Core, RIKEN Brain Science Institute, ²Graduate School of Science and Engineering, Saitama University

The dendritic arborization sensory neurons of the Drosophila larval peripheral nervous system are useful models to elucidate both general and neuron class-specific mechanisms of neuron differentiation. We present a practical guide to generate and analyze dendritic arborization neuron genetic mosaics.

Neuroscience

Immunohistological Labeling of Microtubules in Sensory Neuron Dendrites, Tracheae, and Muscles in the Drosophila Larva Body Wall

Cagri Yalgin^1,2, M. Rezaul Karim^1,2, Adrian W. Moore¹

¹Disease Mechanism Research Core, RIKEN Brain Science Institute, ²Graduate School of Science and Engineering, Saitama University

To understand how complex cell shapes, such as neuronal dendrites, are achieved during development, it is important to be able to accurately assay microtubule organization. Here we describe a robust immunohistological labeling method to examine microtubule organization of dendritic arborization neuron sensory dendrites, trachea, muscle, and other Drosophila larva body wall tissues.

Neuroscience

Whole Mount Immunolabeling of Olfactory Receptor Neurons in the Drosophila Antenna

M. Rezaul Karim¹, Keita Endo², Adrian W Moore³, Hiroaki Taniguchi¹

¹Laboratory for Genetic Code, Graduate School of Life and Medical Sciences, Doshisha University, ²Laboratory for Circuit Mechanisms of Sensory Perception, RIKEN Brain Science Institute, ³Disease Mechanism Research Core, RIKEN Brain Science Institute

Herein we describe the process of whole mount immunostaining of Drosophila antennae, which enables us to better understand the molecular mechanisms involved in the diversification of olfactory receptor neurons (ORN)s.

Developmental Biology

Imaging of Cell Shape Alteration and Cell Movement in Drosophila Gastrulation Using DE-cadherin Reporter Transgenic Flies

M. Rezaul Karim¹, Tomohiro Haruta², Taro Matsumoto³, Hiroki Oda², Hiroaki Taniguchi^3,4,5

¹Biotechnology and Genetic Engineering Department, Jahangirnagar University, ²JT Biohistory Research Hall, ³Division of Cell Regeneration and Transplantation, School of Medicine, Nihon University, ⁴Graduate School of Life and Medical Sciences, Doshisha University, ⁵Institute of Genetics and Animal Breeding of the Polish Academy of Sciences

Herein we describe a procedure to capture live images of Drosophila gastrulation. This has enabled us to better understand the apical constriction involved in early development and further analyze mechanisms governing cellular movements during tissue structure modification.

Biochemistry

A Protein Preparation Method for the High-throughput Identification of Proteins Interacting with a Nuclear Cofactor Using LC-MS/MS Analysis

Megumi Tsuchiya^*1, M. Rezaul Karim^*2, Taro Matsumoto³, Hidesato Ogawa¹, Hiroaki Taniguchi^3,4,5

¹Graduate School of Frontier Biosciences, Osaka University, ²Department of Biotechnology and Genetic Engineering, Jahangirnagar University, ³Division of Cell Regeneration and Transplantation, School of Medicine, Nihon University, ⁴Institute of Genetics and Animal Breeding of the Polish Academy of Sciences, ⁵Graduate School of Life and Medical Sciences, Doshisha University

We have established a method for the purification of coregulatory interaction proteins using the LC-MS/MS system.

Biology

Using a Whole-mount Immunohistochemical Method to Study the Innervation of the Biliary Tract in Suncus murinus

Ke Ren¹, Yidan Dai¹, Kai Yi², Masanobu Kinoshita¹, Masahiro Itoh², Ichiro Sakata³, Takafumi Sakai³, Shuang-Qin Yi¹

¹Department of Frontier Health Sciences, Graduate School of Human Health Sciences, Tokyo Metropolitan University, ²Department of Anatomy, Tokyo Medical University, ³Area of Regulatory Biology, Division of Life Science, Graduate School of Science and Engineering, Saitama University

A whole-mount immunohistochemical approach, to visualize neurofilament protein expression in the extrahepatic biliary tract in Suncus murinus. is presented here. This protocol can be used to analyze the innervation of all visceral organs in S. murinus or other species.

Biology

Real-time In Vivo Recording of Arabidopsis Calcium Signals During Insect Feeding Using a Fluorescent Biosensor

Thomas R. Vincent¹, James Canham¹, Masatsugu Toyota^2,3,4, Marieta Avramova¹, Sam T. Mugford⁵, Simon Gilroy², Anthony J. Miller¹, Saskia Hogenhout⁵, Dale Sanders¹

¹Department of Metabolic Biology, John Innes Centre, Norwich Research Park, ²Department of Botany, University of Wisconsin, Madison, ³Department of Biochemistry and Molecular Biology, Saitama University, ⁴Precursory Research for Embryonic Science and Technology (PRESTO), Japan Science and Technology Agency (JST), ⁵Department of Cell and Developmental Biology, John Innes Centre, Norwich Research Park

This protocol outlines a simple method for analyzing calcium signals in plants generated by feeding hemipteran insects, such as aphids. Arabidopsis thaliana transformed with the GFP calcium biosensor GCaMP3 allow for the real-time in vivo imaging of calcium dynamics with a high temporal and spatial resolution.

Biology

Visualization of DNA Compaction in Cyanobacteria by High-voltage Cryo-electron Tomography

Kazuyoshi Murata¹, Yasuko Kaneko²

¹National Institute for Physiological Sciences, ²Graduate School of Science and Engineering, Saitama University

This protocol describes how to visualize the transient DNA compaction in cyanobacteria. Synchronous cultivation, monitoring by fluorescence microscopy, rapid freezing, and high voltage cryo-electron tomography are used. A protocol for these methodologies is presented, and future applications and developments are discussed.

JoVE Journal

Wide-Field, Real-Time Imaging of Local and Systemic Wound Signals in Arabidopsis

Takuya Uemura¹, Jiaqi Wang¹, Yuri Aratani¹, Simon Gilroy², Masatsugu Toyota^1,2

¹Department of Biochemistry and Molecular Biology, Saitama University, ²Department of Botany, University of Wisconsin

Extracellular glutamate-triggered systemic calcium signaling is critical for the induction of plant defense responses to mechanical wounding and herbivore attack in plants. This article describes a method to visualize the spatial and temporal dynamics of both these factors using Arabidopsis thaliana plants expressing calcium- and glutamate-sensitive fluorescent biosensors.