Name: the power of simplicity: sea urchin embryos as in vivo developmental models for studying complex cell-to-cell signaling network interactions
Uploaded: 2017-02-16T14:00:00
Description: Watch this Scientific Journal Video about The Power of Simplicity: Sea Urchin Embryos as in Vivo Developmental Models for Studying Complex Cell-to-cell Signaling Network Interactions at JoVE.com

We would like to thank Dr. Robert Angerer for his careful reading and editing of the manuscript. NIH R15HD088272-01 as well as the Office of Research and Development, and Department of Biological Sciences at Mississippi State University provided support for this project to RCR.

1.高スループットモルホリノデザイン戦略<ol><li>目的の遺伝子（複数可）（ 例えば 、候補遺伝子アプローチ、シス調節分析、RNAseqおよび/またはプロテオミクス差動画面）を特定します。 </li><li>頻繁に更新されるウェブサイト上で利用可能なゲノム、トランスクリプトーム、および遺伝子発現データを使用してください（ 例えば SpBase http://www.echinobase.org 20と<...

<ol>
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ここで紹介する方法は..多くのラボが分析する早期ウニ開発時に同様のアッセイを使用している基本的な発達のメカニズムを支配するシグナル伝達経路とGRNsを理解するために、脊椎動物未満ゲノムおよび形態学的な複雑さに胚を使用してのパワーを示している例です。他の細胞運命指定イベント（ 例えば、ノッチ、ヘッジホッグ、TGF-βの、及びFGFシグナリング）に関与する経路<sup clas...

遺伝子制御ネットワーク（GRNs）とシグナル伝達経路は、成体動物の体の計画を構築するために使用される胚発生の間の遺伝子の空間的および時間的発現を確立します。細胞から細胞へのシグナル伝達経路は、細胞が通信する手段を提供し、これらの調節ネットワークの重要な構成要素です。これらの細胞の相互作用は、胚発生1、2中の様々な地域でとの間の規制および分化遺伝子の発現を確立し、改良します。分泌された細胞外調節因子（リガンド、アンタゴニスト）、受容体および共受容体の間の相互作用は、シグナル伝達経路の活性を制御します。細胞内分子の品揃えは、遺伝子発現の変化、分割、および/または細胞の形で得られたこれらの入力を伝達します。主要な経路で細胞外および細胞内のレベルで使用する鍵分子の多くがあるが既知の、個々のシグナル伝達経路の複雑さに大部分の不完全な知識です。加えて、異なるシグナル伝達経路は、多くの場合、細胞外、細胞内での正または負のいずれかで互いに相互作用し、転写レベル3、4、5、6。重要なことには、シグナル伝達経路のコアコンポーネントは、非常にすべての後生動物種で保存され、特に密接に関連門の生物を比較する場合、著しく、主要なシグナル伝達経路の大部分は、多くの場合、多くの種において類似の発達機能を実行7、8、9、 10、11。 開発中にシグナル伝達の研究では、任意の生物で困難な作業であり、そしてそこに脊椎動物では、大きな可能性リガンドと受容体/共変調器の相互作用の数、細胞内伝達分子、ならびにがあります1）：ほとんどの新口動物モデル（脊椎動物、無脊椎動物脊索動物、半索動物、および棘皮動物）でシグナル伝達経路を研究するためのいくつかの重要な課題がありますゲノム12の複雑さに起因する異なるシグナル伝達経路の間の潜在的な相互作用、13、14; 2）脊椎動物の複雑な形態および形態形成の動きは、多くの場合、それはより困難でかつシグナル伝達経路間の機能的相互作用を解釈することを可能にします。 3）ほとんどの非棘皮動物無脊椎動物の新口動物モデル種における分析は、いくつかのホヤ種15、16を除いて妊娠の短いウィンドウによって制限されています。 ザウニ胚は、上記制限のいくつかを持っており、 生体内でのシグナル伝達経路の詳細な分析を行うために多くのユニークな資質を提供しています。これらは、以下のものが挙げられる：1）ウニゲノムの相対的なシンプルさが大幅に可能なリガンド、受容体/共受容体と細胞内伝達分子の数を減少させる17の相互対話します。 2）胚葉および主要な胚軸の仕様およびパターニングを制御GRNsはよく信号18、19セル受信/地域の規制状況の理解を助ける、ウニ胚で確立されています。胚は、その形態を分析するのが容易である単層上皮で構成されたとき3）多くのシグナル伝達経路は、早期に切断し、原腸ステージ間で研究することができます。 4）分子が関与しますウニ内シグナル伝達経路中のdは、容易に操作されています。 5）多くのウニは、10〜11ヶ月〜1年（ 例えばStrongylocentrotusのpurpuratusとLytechinusのvariegatus）のために妊娠しています。 ここでは、体系的かつ効率的に指定し、ウニ胚のパターン領土は、いくつかの無脊椎動物モデル系が複雑な分子メカニズムの研究に提供する利点を説明するためにシグナル伝達経路の構成要素を特徴づけるための方法を提示します。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Translational-blocking morpholino and/or splice-blocking morpholino</td>
			<td>Gene Tools LLC</td>
			<td>Customized</td>
			<td>More information at www.gene-tools.com</td>
		</tr>
		<tr>
			<td>Glycerol</td>
			<td>Invitrogen</td>
			<td>15514-011</td>
			<td></td>
		</tr>
		<tr>
			<td>FITC (dextran fluorescein isothiocyanate)</td>
			<td>Invitrogen, Life Technologies</td>
			<td>D1821</td>
			<td>Make 25mg/mL stock solution</td>
		</tr>
		<tr>
			<td>Paraformaldehyde 16% solution EM Grade</td>
			<td>Electron Microscopy Sciences</td>
			<td>15710</td>
			<td></td>
		</tr>
		<tr>
			<td>MOPS</td>
			<td>Sigma Aldrich</td>
			<td>M1254-250G</td>
			<td></td>
		</tr>
		<tr>
			<td>Tween-20</td>
			<td>Sigma Aldrich</td>
			<td>23336-0010</td>
			<td></td>
		</tr>
		<tr>
			<td>Formamide</td>
			<td>Sigma Aldrich</td>
			<td>47671-1L-F</td>
			<td></td>
		</tr>
		<tr>
			<td>Yeast tRNA</td>
			<td>Invitrogen</td>
			<td>15401-029</td>
			<td></td>
		</tr>
		<tr>
			<td>Normal Goat Serum</td>
			<td>Sigma Aldrich</td>
			<td>G9023-10mL</td>
			<td></td>
		</tr>
		<tr>
			<td>Alkaline Phosphatase-conjugated anti-digoxigenin antibody</td>
			<td>Roche</td>
			<td>11 093 274 910</td>
			<td></td>
		</tr>
		<tr>
			<td>Tetramisole hydrochloride (levamisole)</td>
			<td>Sigma Aldrich</td>
			<td>L9756-5G</td>
			<td></td>
		</tr>
		<tr>
			<td>Tris Base UltraPure</td>
			<td>Research Products Internationall Corp</td>
			<td>56-40-6</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium Chloride</td>
			<td>Fisher Scientific</td>
			<td>BP358-10</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnesium chloride</td>
			<td>Sigma Aldrich</td>
			<td>7786-30-3</td>
			<td></td>
		</tr>
		<tr>
			<td>BCIP (5-Bromo-4-Chloro-3-indolyl-phosphate</td>
			<td>Roche</td>
			<td>11 383 221 001</td>
			<td></td>
		</tr>
		<tr>
			<td>4 Nitro blue tetrazolium chloride (NBT)</td>
			<td>Roche</td>
			<td>11 383 213 001</td>
			<td></td>
		</tr>
		<tr>
			<td>Dimethyl Formamide</td>
			<td>Sigma Aldrich</td>
			<td>D4551-500mL</td>
			<td></td>
		</tr>
		<tr>
			<td>Potassium Chloride</td>
			<td>Sigma Aldrich</td>
			<td>P9541-5KG</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium Bicarbonate</td>
			<td>Sigma Aldrich</td>
			<td>S5761-500G</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnesium Sulfate</td>
			<td>Sigma Aldrich</td>
			<td>M7506-2KG</td>
			<td></td>
		</tr>
		<tr>
			<td>Calcium Chloride</td>
			<td>Sigma Aldrich</td>
			<td>C1016-500G</td>
			<td></td>
		</tr>
	</tbody>
</table>

the power of simplicity: sea urchin embryos as in vivo developmental models for studying complex cell-to-cell signaling network interactions

Remarkably few cell-to-cell signal transduction pathways are necessary during embryonic development to generate the large variety of cell types and tissues in the adult body form. Yet, each year more components of individual signaling pathways are discovered, and studies indicate that depending on the context there is significant cross-talk among most of these pathways. This complexity makes studying cell-to-cell signaling in any in vivo developmental model system a difficult task. In addition, efficient functional analyses are required to characterize molecules associated with signaling pathways identified from the large data sets generated by next generation differential screens. Here, we illustrate a straightforward method to efficiently identify components of signal transduction pathways governing cell fate and axis specification in sea urchin embryos. The genomic and morphological simplicity of embryos similar to those of the sea urchin make them powerful in vivo developmental models for understanding complex signaling interactions. The methodology described here can be used as a template for identifying novel signal transduction molecules in individual pathways as well as the interactions among the molecules in the various pathways in many other organisms.

ウニ胚では、3つの異なるWntシグナル支店（のWnt /βカテニンは、Wnt / JNK、およびWnt / PKC）4、25相互作用は前後（AP）パターニングを支配するWntシグナル伝達ネットワークを形成することが示されています。これらのシグナル伝達イベントの中で最も重要な結果の一つは、初期広く発現し、前方神経外胚葉（ANE）GRNは、原腸形?...

Watch this Scientific Journal Video about The Power of Simplicity: Sea Urchin Embryos as in Vivo Developmental Models for Studying Complex Cell-to-cell Signaling Network Interactions at JoVE.com

The Power of Simplicity: Sea Urchin Embryos as in Vivo Developmental Models for Studying Complex Cell-to-cell Signaling Network Interactions

このビデオの記事では、体系的かつ効率的に多くの無脊椎動物の胚では、複雑なシグナル伝達経路および制御ネットワークの構成要素を特徴付けるために使用することができるin vivoでの方法論に簡単な詳細を示します。

シンプルの力：ウニ胚として<em&gt; in vivoで</em&gt;複雑な細胞間シグナル伝達ネットワークの相互作用を研究するための発達モデル

Remarkably few cell-to-cell signal transduction pathways are necessary during embryonic development to generate the large variety of cell types and ...

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The Power of Simplicity: Sea Urchin Embryos as in Vivo Developmental Models for Studying Complex Cell-to-cell Signaling Network Interactions

Unit 1

Principles of Cell Signaling

SCIENTISTS IN ACTION

Protocols for Obtaining Zygotic and Somatic Embryos for Studying the Regulation of Early Embryo Development in the Model Legume Medicago truncatula

Protocols for Obtaining Zygotic and Somatic Embryos for Studying the Regulation of Early Embryo Development in the Model Legume Medicago truncatula

モデルマメに初期胚発生の調節を研究するための接合体と体細胞胚を得るためのプロトコル<em&gt;のMedicago truncatula</em

Early embryogenesis starting from a single cell zygote goes through rapid cell division and morphogenesis, and is morphologically characterized by ...

Characterization of Thymic Settling Progenitors in the Mouse Embryo Using In Vivo and In Vitro Assays

Characterization of Thymic Settling Progenitors in the Mouse Embryo Using In Vivo and In Vitro Assays

胸腺の特性を用いてマウス胚における前駆細胞をセトリング<em&gt;インビボ</em&gt;と<em&gt;インビトロ</em&gt;アッセイ

Characterizing thymic settling progenitors is important to understand the pre-thymic stages of T cell development, essential to devise strategies for T ...

Studying Wnt Signaling During Patterning of Conducting Airways

誘導気道のパターニング時にWntシグナル伝達を学びます

Wnt signaling pathways play critical roles during development of the respiratory tract. Defining precise mechanisms of differentiation and morphogenesis ...

Inducing Complete Polyp Regeneration from the Aboral Physa of the Starlet Sea Anemone Nematostella vectensis

Inducing Complete Polyp Regeneration from the Aboral Physa of the Starlet Sea Anemone Nematostella vectensis

スターレットイソギンチャクの尾方Physaから完全ポリープ再生を誘導<em&gt; Nematostella vectensis</em

Cnidarians, and specifically Hydra, were the first animals shown to regenerate damaged or severed structures, and indeed such studies arguably launched ...

Drug Treatment and In Vivo Imaging of Osteoblast-Osteoclast Interactions in a Medaka Fish Osteoporosis Model

Drug Treatment and In Vivo Imaging of Osteoblast-Osteoclast Interactions in a Medaka Fish Osteoporosis Model

薬物治療と<em&gt;インビボ</em&gt;メダカ骨粗鬆症モデルにおける骨芽細胞破骨細胞の相互作用のイメージング

Bone-forming osteoblasts interact with bone-resorbing osteoclasts to coordinate the turnover of bone matrix and to control skeletal homeostasis. Medaka ...

In Vivo Imaging of Transgenic Gene Expression in Individual Retinal Progenitors in Chimeric Zebrafish Embryos to Study Cell Nonautonomous Influences

In Vivo Imaging of Transgenic Gene Expression in Individual Retinal Progenitors in Chimeric Zebrafish Embryos to Study Cell Nonautonomous Influences

細胞非自律影響を研究するためのキメラゼブラフィッシュ胚における個々の網膜前駆細胞におけるトランスジェニック遺伝子発現のin vivoイメージング

The genetic and technical strengths have made the zebrafish vertebrate a key model organism in which the consequences of gene manipulations can be traced ...

Live Imaging of Mouse Secondary Palate Fusion

マウス二次口蓋融合のライブイメージング

The fusion of the secondary palatal shelves to form the intact secondary palate is a key process in mammalian development and its disruption can lead to ...

Visualizing the Node and Notochordal Plate In Gastrulating Mouse Embryos Using Scanning Electron Microscopy and Whole Mount Immunofluorescence

ノードと走査型電子顕微鏡と蛍光抗体法マウント全体を使用して Gastrulating のマウス胚における脊索板を可視化

The post-implantation mouse embryo undergoes major shape changes after the initiation of gastrulation and morphogenesis. A hallmark of morphogenesis is ...

Assessment of Oxidative Damage in the Primary Mouse Ocular Surface Cells/Stem Cells in Response to Ultraviolet-C (UV-C) Damage

Assessment of Oxidative Damage in the Primary Mouse Ocular Surface Cells/Stem Cells in Response to Ultraviolet-C UV-C Damage

紫外C(UV-C)損傷に対する主なマウス眼表面細胞/幹細胞における酸化的損傷の評価

The ocular surface is subjected to regular wear and tear due to various environmental factors. Exposure to UV-C radiation constitutes an occupational ...

A Layered Mounting Method for Extended Time-Lapse Confocal Microscopy of Whole Zebrafish Embryos

ゼブラフィッシュ胚全体の延長タイムラプス共焦点顕微鏡法の層状取り付け法

Dynamics of development can be followed by confocal time-lapse microscopy of live transgenic zebrafish embryos expressing fluorescence in specific tissues ...