ブラッドフォード法が良く使われています。
原著論文
A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding Marion M. Bradford Analytical Biochemistry Volume 72, Issues 1–2, 7 May 1976, Pages 248-254 https://www.sciencedirect.com/science/article/abs/pii/0003269776905273 本文有料
パラグラフライティングという言葉はすっかり巷に流布していますが、実際にパラグラフライティングが何かを知っている人はあまりいないのではないでしょうか。自分も、せいぜいトピックセンテンスをパラグラフの最初に書くやつね？くらいの認識しかありませんでした。しかし、図書館で「理系のパラグラフライティング」（羊土社）という本を見つけて開いてみたら、もっと具体的で系統だった説明がありました。その中の内容をネタに、自分なりに説明してみます。
パラグラフライティングは、お作法です。決め事があります。まず、パラグラフの第1文は、トピックセンテンスと呼ばれるものを書きます。トピックセンテンスとは、トピック（パラグラフの主題）があり、それがどうした？の部分「コントローリングアイデア」が続いて、1文になります。2文目からは、そのトピックセンテンスの主張をサポートする、サポーティングセンテンスを書いていきます。サポートの中身としては、①さらに詳細な説明、②具体例、③理由や根拠が書かれるのが一般的です。そしてパラグラフの最後は、コンクルーディングセンテンスで締めくくります。コンクルーディングセンテンスは、書かれないこともあるようです。
作法としてはこれだけなので話は単純なはずですが、「知っている」ことと「できる」こととは大違いで、実際に書いてみるとこの作法に従わない文章を平気で書いてしまうものです。
ありがちな失敗としては、まず「トピックセンテンス」がパラグラフの中に存在しないということがあります。自分がトピックセンテンスのつもりで書いていても、そうなってないようというわけです。主張＋サポート（具体例）の形があることからわかるように、トピックセンテンスの主張にはある程度の「抽象性」が要求されます。最初に抽象的に書くからこそ、そのあとで具体例が書けるのです。
サポーティングセンテンスに関してもアリがちな失敗がいくつかあります。一つ目は、トピックをサポートしない情報を書いてしまう、つまり、不要なことを書いてしまうことです。知っていることは何でもつい書いておきたくなるのが人情ですが、それをやると、締まりのない文章になります。2つ目の失敗は、必要なことを書かないことです。主張をする以上、その主張が抽象的なのであれば具体例が必要ですし、その主張が直ちに読者に受入れられないようなことであれば、根拠を示す必要があります。
コンクルーディングセンテンスを書くときの失敗例としては、トピックセンテンスで述べた以上のことを書いてしまうことなどが挙げられます。最後になって突然新たな主張を始めたり、主張を拡大してみたりするのはダメよというわけです。コンクルーディングセンテンスは、内容としてはトピックセンテンスと完全に一致していないといけません。かといってトピックセンテンスをコピペしてパラグラフの最後に持って来るのも芸がありませんので、言葉を変えて表現する必要があります。
この本の教えだと第1文は必ずトピックセンテンスでありコントローリングアイデアを含めるようにとのことです。ただ現実的には、トピックがあまり読者に馴染が無い場合にはトピックの背景や詳細を説明してから、主張を書くということは普通にありますので、あまり「型にはめること」を優先してしまうと本末転倒になるのではないかと思いました。一番優先すべきは、読みやすいこと、理解しやすいこと、文章に自然な流れがあることでしょう。
理系の人に役立つパラグラフライティングの本は今まで類書を見たことがなかったので、この本はなかなか良い教材になると思います。また、学生に教える立ち場の人にとっても、演習のために使う教材、指定教科書・指定参考書になるのではないでしょうか。
パラグラフの書き方をマスターしただけでは実は不十分で、パラグラフとパラグラフのつながりも必要です。多くの場合に、内容によってつながっていくと思います。
SNSを見ていたら、脳外科医”竹田くん”の話題が目に留まりました。知らない人のために言っておくと「竹田」は漫画の主人公の名前であって、モデルと噂されている実在する人物の実名ではありません。また実在する人物の氏名と同姓同名の医師は日本に複数おられるようですが、問題視されている医師は2024年現在、吹田市の徳洲会病院の救急科（ER）に常勤医として勤務しているそうです（下の、院長の声明参照）。この事実を知ってしまった以上、吹田市徳洲会病院は、自分だったら絶対に救急車で運ばれたくない病院になってしまいました。
脳外科医 竹田くん あり得ない脳神経外科医 竹田くんの物語（漫画のサイト）
アーカイブ（月）ごとに辿ると第1話から順に読みやすいと思います。アーカイブ2023/01 【第１話】履歴書 【第２話】後継者 【第３話】初登院 【第４話】男の約束 【第５話】脳外チーム 2023 / 2 (34) 2023 / 3 (11) 2023 / 4 (21) 2023 / 5 (27) 2023 / 6 (24) 2023 / 7 (15) 【第１４２話】霧（最終話 2023年7月22日投稿）
医道審議会は、罰金刑以上が確定した医師や看護師など医療職の犯罪者に行政処分を下す厚生労働省の審議会です。
懲役や罰金など、犯罪のレベルによって、刑罰の重さが変わるのと同じです。過去の事例から、児童福祉法違反は重く、児童ポルノ禁止法違反は軽く、といった処分になります。— MOMA (@msnNightingale) November 21, 2024
本当にその通り。
厚労省やその下にある医道審議会が無能過ぎる。
医道審が動くのは刑事処分の後追いだけ。
自分らでは全く判断しない無能組織。 https://t.co/I9eM0cTqxq— niko＠医療過誤（医療ミス）真実を知りたい (@tokitako73) November 20, 2024
脳外科医竹田くんの件、厚労省の医道審議会が機能してないのが諸悪の根源なのよね。
医師免許取り上げがほぼない。— ゆな先生 (@JapanTank) November 20, 2024
脳外科医の竹田くんが話題だけど、医師免許の扱いを判断する医道審議会が腰抜けだから期待できない。東京で7人も女性を盗撮し有罪が確定した医者を、たった7カ月で現場復帰させる程度の倫理観しかない。 https://t.co/6rVyZ7Pxls
— みさき (@T_Tmisaki) November 20, 2024
「医道審議会(医道分科会)」が竹田くんを処分できていなかったことが最大の問題点なんじゃないかな？
せめて世にはびこる反ワク医師を処分できるくらいには組織改編して欲しい。 https://t.co/fgMX5lxuH3— GikuGiku (@gikugikuhiiiii) November 20, 2024
▼追記: 吹田徳洲会の職員さんより頂いたタレコミ
・本日、週刊紙の記事について高橋俊樹院長が職員向けに説明会を開いた
・全体的にA氏を擁護する内容■説明会要約
・記事にはデタラメばかりが書いてある
・A氏が起こした医療過誤はすべて過去の話、本人は反省しているので掘り返すべきではない…— 滝沢ガレソ (@tkzwgrs) May 8, 2024
世間から問題視されている医師は、現在、吹田徳洲会病院の救急科で常勤の職を得て働いているそうです。下の院長の声明を読む限り、院長もこの医師によるトラブルを認めており、週刊誌への告発内容には信憑性があるようです。
一部週刊誌に掲載された記事について 先日、某週刊誌に掲載された当院救急部門の ER 医師に関する記事について、これまで の経緯と現状について述べさせていただきます。 雑誌社から質問状が届きインタビューに応じました。質問された各事象については医療 過誤と言えるものが一件もないことを医学的観点から丁寧に説明し正直かつ誠実に回答し ました。しかし、記事にはほとんど反映されず、昨年夏から冬にかけて報告された内容を いたずらに曲解・誇張し、誤った誹謗中傷の情報まで記載されていました。的外れで、事 実を捻じ曲げ、悪い評判を煽るだけの今回の週刊誌編集部に対しては断固として抗議を行 うつもりです。 該当医師の ER での診療態度やスタッフとの付き合い方に問題があったのは事実です。 病院として、注意や警告も含めて何度も指導してきたつもりでしたが、その効果が出るの に予想以上に時間がかかり、ER現場のスタッフには思いがけないストレスをかけることに なってしまいました。これが今回の週刊誌へのリークに至った理由です。紆余曲折があり ましたが、ここに来て当該医師も診療態度を改め、スタッフもチーム医療を最優先とした ＥＲ運営ができるようになりました。当該医師の ER 医師としての診療能力は ER と連携す る各診療科の責任者に聞いても評価されるようになってきており、頑張ってくれている、 との声も聞こえてくるようになりました。さらに一般外来に専門医の診察を要する重症患 者さんが来られた場合にも迅速かつ的確に ER で対応できる体制となっています。 当院の ER で定期的に勤務していただいている非常勤の先生方は診療経験豊かなベテラ ン医師が多く、若い先生方も大学医局からの派遣で熱心な医師ばかりです。 常勤医としての自覚をしっかりと持つようになった当該医師が加わって、今後の当院の ER 診療は地域の皆さんの期待に今まで以上に応えることができるものと考えています。 今後ともどうかよろしくお願い申し上げます。 2024 年 5 月 8 日 吹田徳洲会病院 病院長 高橋 俊樹 https://www.suita.tokushukai.or.jp/wp-content/uploads/2024/05/2208aa85824a8082645325cc76898fbd-1.pdf
赤穂市民病院 脳神経外科における医療事故に関する記者会見（2022年6月28日） 赤穂民報チャンネル チャンネル登録者数 483人 (2:00:58)
神戸徳洲会の件は循内で「竹田くん」は脳外やからおそらく別人
「竹田くん」もさすがにPCIはできんやろ https://t.co/DFmPvVGKaz— さぼりーまん (@75QUWuAmJJ84265) March 18, 2024
ヒスチジンは生理的なpH＝7.4ではプロトン化されていませんが、弱酸性になるとプロトン化されています。
The imidazole ring in histidine exhibits a pKa value within the pH range from 5.5 to 7.4); at pH 5.0, the imidazole ring is protonated and positively charged, whereas, at pH 7.0, it is electrically neutral. https://www.jstage.jst.go.jp/article/bpb/44/5/44_b20-01013/_html/-char/ja
研究セミナーを聞いていたら、ヒスチジンのプロトン化がpHセンサーとして働くという例が示されていて、「へぇ～」と思いました。生き物はうまくできていますね。ヒスチジンのプロトン化は、pHセンサーとして働く意外にも、様々な重要な役割がありました。
酵素の活性中心に存在するヒスチジンは、pHに応じてプロトン化・脱プロトン化しやすい性質を持ち、酸-塩基触媒として機能します。 例: セリンプロテアーゼ（例: キモトリプシン）では、ヒスチジンが触媒三重体の一部として、基質の加水分解に関与します。 **カルボン酸脱水酵素（炭酸脱水酵素）**では、ヒスチジンがH⁺の授受を媒介して反応を促進します。 意義: ヒスチジン残基のプロトン化状態が変化することで、酵素反応の速度や特異性が制御されます。
ヒスチジンは、側鎖のpKaが約6.0と生理的pH（7.2〜7.4）に近いため、小さなpH変化に敏感に反応します。 例: イオンチャネルや輸送体タンパク質では、ヒスチジンがpH変化を感知して活性を調節します。 **pH感受性Gタンパク質共役受容体（GPCRs）**では、ヒスチジン残基が細胞外のpH変化に応答します。 意義: 組織や細胞環境の酸塩基平衡を感知することで、生体の恒常性維持に寄与します。
ヒスチジンは、プロトン化状態によって金属イオン（Zn²⁺, Cu²⁺, Fe²⁺など）への親和性が変化し、タンパク質の構造や機能に影響を与えます。 例: ヘモグロビンでは、ヒスチジンが鉄（Fe²⁺）と結合し、酸素結合に寄与します。 メタロ酵素（例: カルボン酸脱水酵素やスーパーオキシドジスムターゼ）では、金属イオンとの結合を通じて触媒活性を発揮します。 意義: 金属イオンの結合や解離を調節し、酵素の触媒機能や基質特異性を制御します。
ヒスチジンのプロトン化状態が変化すると、分子間の静電相互作用や水素結合の形成が変化し、タンパク質間相互作用を調節します。 例: シグナル伝達経路の中で、タンパク質の結合部位のヒスチジンがプロトン化状態を変えることで結合強度が調整されます。 ヒスチジンリン酸化（プロカリオットでよく見られる）は、細胞内シグナル伝達に関与します。 意義: タンパク質の動的な挙動や複合体形成に重要です。
プロトン化されたヒスチジン残基は、タンパク質の三次構造や四次構造を安定化または不安定化する場合があります。 例えば、酸性環境下では、ヒスチジンのプロトン化により静電相互作用が変化し、タンパク質のフォールディングやアンフォールディングに影響を与えます。 pHに依存したタンパク質の構造変化や機能調節に寄与しするという意義があります。
ヒスチジン残基がプロトンの供与体または受容体として働き、細胞内外のpH調節に関与します。 例: ヘモグロビンのボーア効果では、ヒスチジンのプロトン化状態が酸素解離能を調節します。 意義: 酸素輸送や代謝調節において重要な役割を果たします。
総説論文（review article, review paper）には、目的や形式によっていくつかの種類があります。総説論文を書く際には、そもそも自分はどの種類の総説論文を書こうとしているのかを決める必要があります。逆に自分が総説論文を読む場合には、その総説論文がどの種類に該当するものなのかを素早く見抜く必要があります。
総説論文の種類に関して、代表的なものを見てみましょう。
特定のトピックに関する研究を包括的にまとめて、そのテーマの全体像を提示してみせるようなレビューはNarrative Review（ナラティブレビュー）と呼ばれます。ナラティブレビューの特徴としては、 執筆者の独自の視点に基づいているということが上げられます。つまり、ナラティブレビューを書く人が自分で問題意識を持ち、問題設定をしてそれに基づいて文献を調べてまとめるというものです。ナラティブレビューは主観的な要素が強く、 レビュー論文を書くにあたって文献をどのようにして系統的に検索したかという方法を示すことは要求されていません。注目した論文の選び方にそもそも主観が入るというわけです。
Systematic Review（系統的レビュー）というのは、特定の研究課題について、事前に定めた方法論に基づき、系統的に客観性をもった手順で文献を収集し、評価し分析してかかれたレビュー論文です。その特徴:としては、明確なリサーチクエスチョンが設定されます。その際には、いわゆるPICOなどのフレームワークが利用されます。また、文献検索のプロセスは詳細に記載されるのが普通です。
Meta-Analysis（メタアナリシス）は文字通り「分析」であり、系統的レビューの一部として行われることが多いです。全ての系統的レビュー論文がメタアナリシスを含むというわけではありません。逆に、メタアナリシスが必ず統計的レビューの一部とうわけでもなく、すでに統計的レビュー論文が存在する場合に、その統計的レビュー論文に基いてメタアナリシスを実施して、「メタアナリシス論文」として発表されることもあります。
統計的手法を用いて複数の研究結果を統合・解析するレビューであるという特徴があります。 とくに、定量的な結果を導き出すことが目的です。異なる研究論文を比較することにより、一貫性や総合的な効果を検討します。
Scoping Review（スコーピングレビュー）は、設定したテーマに関して全体的な範囲を探索し、主要な概念、証拠の種類、研究ギャップなどを特定するためのレビュー論文です。 特徴としては、メタアナリシスにみられるような厳密な評価や統合を必ずしも目的としないということがあります。 例えば、「人工知能を用いた医療診断に関する研究の現状」といったテーマがスコーピングレビューとしては考えられます。
スコーピングレビューとナラティブレビューの共通点として、どちらも広範なトピックを扱い、特定のテーマについて包括的に情報を収集し、全体像を示すことを目的としているということがあります。また、メタアナリシスのような厳密な統計解析を行わず、定量的な統合は行わないこと、質的な情報を主に扱うことにおいても共通しています。系統的な手法が必須とはされておらず、: スコーピングレビューもナラティブレビューも、方法論的には柔軟なアプローチがとられます。
スコーピングレビューがナラティブレビューと異なる点はいくつかあります。まず、目的が明確に異なります。ナラティブレビューでは、知識の整理やトピックの概要を提供するという目的があります。執筆者の主観に基づき、情報の解釈や選択が行われます。それに対して、スコーピングレビューではトピックの範囲をマッピングすることに焦点を当てます。「何がわかっているか」「研究のギャップはどこか」を明らかにすることはスコーピングレビューの目的です。スコーピングレビューは、ナラティブレビューに比べて、もっと構造化された方法論を用いることが求められます。またその方法論の記載すんわち、透明性も要求されます。スコーピングレビューでは、系統的レビューほど厳密ではないものの、文献検索の手順や選定基準を明示することが求められます。
スコーピングレビューは、研究分野がまだ発展途上であり、系統的レビューを行うのに十分なエビデンスが揃っていない場合に特に有用とされます。それに対して、ナラティブレビューは、発展段階に関わらず幅広いテーマで行われることが多いといえます。
スコーピングレビューもナラティブレビューも、とられる方法論に柔軟性があること、定量的な分析を伴わないこと、研究のギャップの特定や広範な概要を示すことなどはナラティブレビューの目的と重なることから、スコーピングレビューをナラティブレビューの一形態とみなす場合もあり得ます。
Critical Review（クリティカルレビュー）は、執筆者が選んだトピックに関連する文献を批判的に評価することにより、現在受け入れられている仮説を見直して、新たな洞察や解釈を提供するレビュー論文です。 クリティカルレビューの特徴は、過去に行われたそれまでの研究に対して批判的に解釈し、研究に関する改善の提案、新たな仮説の可能性の提示を行うものです。クリティカルレビューにおいては、 執筆者の分析力や理論的な視点・考察が重要になります。
ミニレビューは話題を限局し、特定の側面について簡潔にまとめたレビュー論文です。 通常、1,000～3,000語程度の短い記事になります。新規性の高いトピックや急成長している分野で執筆されることが多い種類の論文です。
アンブレラレビューとは、複数の系統的レビューやメタアナリシスを統合し、広範なテーマについて俯瞰的な結論を導き出すレビューです。「レビューのレビュー」ともいえます。そのため、エビデンスレベルが高い論文とされます。
文字通り、特定の研究分野の最前線の進展や知識をまとめたレビュー論文になります。 最新の研究成果に焦点を当てているという特徴があります。 将来の研究課題や展望を示すことが多いです。例えば、「量子コンピューティングの現状と未来」といったテーマがありえます。
ヒストリカルレビューとは、特定のトピック・分野の歴史的な発展を追ったレビュー論文です。 時系列に沿って知識や理論の進展を記述するという特徴があります。 現在の理解に至った歴史的経緯、背景が書かれます。「がん研究の歴史的展望」といったテーマが例としてありえます。
言葉の定義
アセクシュアル（asexual）：If you identify as asexual, you may experience a little sexual attraction or none at all.
アロマアセク：恋愛感情を抱かないアロマンティックと性的欲求を抱かないアセクシュアルの2つのセクシュアリティをもち合わせるセクシュアリティ
バイセクシャル：男性と女性、両方の性に対して恋愛感情を抱いたり性的にも惹かれたりするセクシュアリティ
恋愛的指向（romantic orientation）：どちらの性別に対して恋愛感情をいだくかに関する傾向のこと。
クロス・オリエンテーション（cross orientation）：性的指向と恋愛的指向が一致しないこと
参考
Int J Mol Sci. 2021 Oct 13;22(20):11030. doi: 10.3390/ijms222011030 https://pmc.ncbi.nlm.nih.gov/articles/PMC8538579/
間葉が表皮に働きかけて、表皮の一部がプラコード（原基）に分化します。
09 June 2021 A 4D road map for the formation of hair follicles https://www.nature.com/articles/d41586-021-01482-1
Organogenesis. 2018 Feb 15;14(1):46–63. doi: 10.1080/15476278.2017.1421882 Embryonic skin development and repair https://pmc.ncbi.nlm.nih.gov/articles/PMC6150059/
ChatGPTの解説です。
毛を作る細胞やその構成成分についての具体的な仕組みを以下に整理します。
毛を作る細胞は、主に以下のようなものがあります：
Hair bulb matrix cells are unspecialized epithelial cells that grow the hair follicle and hair:
毛髪の主成分は**ケラチン（Keratin）**です。
毛の色は、**メラニン（Melanin）**という色素によって決まります。
毛包と毛髪の構造や性質は、体の部位によって異なります。
以下の説明もChatGPT4oによるもの。
EDA（Ectodysplasin A）とEDAR（Ectodysplasin A receptor）は、主に外胚葉由来の構造（毛包、汗腺、歯、乳頭など）の形成や分化に関与する重要な分子とその受容体です。これらはTNF（Tumor Necrosis Factor）ファミリーに属し、細胞間のシグナル伝達を介して発生過程を調節します。
以下にEDA/EDARシグナル伝達機構の詳細を説明します。
EDA/EDARシグナルの異常は、さまざまな発生異常を引き起こします。
EDA/EDARは、外胚葉由来の構造形成（毛、汗腺、歯など）において不可欠な役割を果たすシグナル伝達機構です。その異常は発生異常や疾患につながり、これらの分子を標的とした再生医療や治療法の開発が進んでいます。このシグナルの動的な調節が、発生過程における細胞の分化と形態形成を支えています。
MINI REVIEW article Front. Genet., 03 April 2023 Volume 14 – 2023 | https://doi.org/10.3389/fgene.2023.1168538 The EDA/EDAR/NF-κB pathway in non-syndromic tooth agenesis: A genetic perspective https://www.frontiersin.org/journals/genetics/articles/10.3389/fgene.2023.1168538/full
Nature Communications volume 9, Article number: 2333 (2018) Published: 20 June 2018 FGF signalling controls the specification of hair placode-derived SOX9 positive progenitors to Merkel cells
STEM CELLS Tissue-Specific Stem Cells Free Access Concise Review: Wnt Signaling Pathways in Skin Development and Epidermal Stem Cells Anthony Veltri, Christopher Lang, Wen-Hui Lien First published: 19 October 2017 https://doi.org/10.1002/stem.2723 https://stemcellsjournals.onlinelibrary.wiley.com/doi/10.1002/stem.2723
Front Med (Lausanne). 2022 Apr 14;9:863786. doi: 10.3389/fmed.2022.863786 Epithelial-Mesenchymal Interaction in Hair Regeneration and Skin Wound Healing https://pmc.ncbi.nlm.nih.gov/articles/PMC9048199/
（雑誌社の論文もフリーだがその図よりもSEMANTICSCHOLARのウェブサイトの図のほうが高解像度） https://www.semanticscholar.org/paper/Maintaining-Hair-Inductivity-in-Human-Dermal-Cells%3A-Taghiabadi-Nilforoushzadeh/c05f7a947c658f8921ebb08182f27897141f625a/figure/0
皮膚の発生・形成機構に関するChatGPTのまとめ：
真皮の形成は複雑で、多くのシグナル伝達経路が関与しています。以下に、真皮の形成をより詳細に説明し、BMPの発生源、標的、作用、部位ごとの違い、そして真皮の機能（髪の毛の形成や表皮との関係）について整理します。
これにより、体の部位ごとに真皮の起源が異なるため、特性や厚みも部位ごとに変わります。
spondylothoracic dysostosis (STD) (Lavy-Moseley syndrome) とspondylocostal dysostosis (SCD) (Jarcho-Levin syndrome)について。
1938年にジョンズ・ホプキンス病院のSaul JarchoとPaul Levinが、椎骨および肋骨の異常による胸部不全の症例を報告しました。Jarcho-Levin症候群 (SCD)は、Notch経路に関連するDLL3遺伝子の異常が関与しています。罹患は人種を問いません。
1966年にはインディアナ大学のNorman Lavyらが、プエルトリコ出身の家族においてJarchoとLevinが報告した症例と同様の症候群を報告しました。Lavy-Moseley症候群(STD) は、Notch経路の遺伝子であるMESP2遺伝子の異常が関与しています。こちらはプエルトリコ人に多くみられます。
体節の基になる細胞は実は体の一番後ろの尾部 （原始索条または尾芽）から生まれます。そこで細胞が増殖して前へ前へと押し出されていきます。
理科年表Q&A 体節のでき方2008.04.01 【相賀裕美子 国立遺伝学研究所（2008年 4月）】https://official.rikanenpyo.jp/posts/6668
脊椎動物の体節は、吻側側から一つ一つできていきます。この体節ができるメカニズムに関しては、多くのシグナルが関与することが知られています。吻側側ではレチノイン酸が分泌されて、尾側にかけて勾配をつくっています。逆に尾側ではFGFとWNTが分泌されており吻側にかけて濃度勾配ができています。レチノイン酸とFGFは互いに拮抗するように作用します。また、いくもの遺伝子発現が周期的に変動することが知られており、その変化が尾側から吻側へと移動していきます。そして、次に大切になる部分でMesp2遺伝子などが特異的に発現し、新たな体節が形成されます。
体節がひとつずつ付け足されていくという形態学的な変化はわかりやすいですが、体節ができるタイミングとおなじ周期性をもって遺伝子発現が変動しているという発見は、体節形成の研究者にとって大きな驚きだったようです。
ポスドク時代に、体節形成に重要な役割を果たす遺伝子を研究していたときのことだった。目的の遺伝子の発現を、標識したRNAプローブを結合させることで検出したが、何度やっても検出できる検体とできない検体があった。「実験に失敗したものと思っていました。でも、本当は失敗ではなく、全部事実だったんです」。その遺伝子は2時間間隔という発現周期を持っていたのだ。「まさか、そんなにダイナミックな発現のON/OFFが起こっているなんて、当時は考えてもみなかったんです」。自分が出した結果をよく見つめ直し、どこが成功でどこが失敗なのかをしっかり解析することで、初めて次のステップに進むことができるのだ。https://www.nig.ac.jp/nig/ja/research/interviews/faculty-interviews/yumiko-saga
Published: 28 February 2024 Cellular and molecular control of vertebrate somitogenesis Yuchuan Miao & Olivier Pourquié Nature Reviews Molecular Cell Biology volume 25, pages517–533 (2024)
https://www.nature.com/articles/s41580-024-00709-z
下の動画の説明がわかりやすかった。
Somite Formation & 90min clock Crash Biology チャンネル登録者
HEXは体節形成において周期的な遺伝子発現レベルの変動を示す分子の一つです。HEXは転写抑制因子として自分自身の遺伝子発現も抑制します。転写、翻訳、抑制に遅延時間が存在することによりHEXのmRNA量およびタンパク質量が周期的な変動を示すことがモデリングにより示されています（Lewisのカレントバイオロジー論文）。遺伝子の発現が周期的に変動するのは非常に不思議なことですが、その不思議な現象が実は複雑は転写因子のネットワークなどを必要とせず、たった一つの転写抑制因子だけで起こりえるというのもまた非常に面白味があります。
FGF (Fibroblast Growth Factor) does play a role in suppressing new somite formation, particularly in the posterior presomitic mesoderm (PSM). High levels of FGF in the posterior PSM maintain cells in an unsegmented, undifferentiated state. This suppression helps create the gradient required for the orderly, sequential formation of somites as cells move from the posterior to the anterior region of the PSM.
If FGF signaling is experimentally suppressed or reduced:
- Premature Somite Formation: Cells that would normally be held in an unsegmented state in the posterior PSM can undergo premature segmentation. This can lead to the formation of somites before the cells reach the proper position in the anterior PSM, where segmentation is normally coordinated.
- Disruption of Rostrocaudal Polarity: Lowered FGF levels might interfere with the normal rostrocaudal patterning within somites. FGF signaling is also involved in establishing the correct polarity within each somite, so its suppression can cause defects in somite boundary formation and polarity.
- Alteration of Determination Front Position: The “determination front” — the zone where cells transition from unsegmented to segmented states — is influenced by the balance of FGF and RA gradients. Suppressing FGF shifts this determination front posteriorly, resulting in somites forming prematurely, closer to the posterior PSM.
Experimental studies have shown that reducing FGF signaling can indeed lead to the formation of smaller, irregularly patterned somites, as cells segment earlier than intended. This highlights FGF’s role in controlling the timing and spatial pattern of somite formation, ensuring that somites form in an orderly, regulated manner as cells progress through the PSM.
(ChatGPT)
FGF and Wnt signaling are critical for activating and maintaining Tbx6 gene expression in the posterior presomitic mesoderm (PSM) during somitogenesis. Here’s how these pathways regulate Tbx6:
- Activation and Maintenance of Tbx6 Expression: High levels of FGF and Wnt signaling in the posterior PSM provide a supportive environment that promotes Tbx6 expression. These signals maintain Tbx6 at levels necessary for keeping cells in an undifferentiated, mesenchymal state, preparing them for future segmentation as they move toward the anterior.
- Regulation Through Gradients: FGF and Wnt form gradients that are highest in the posterior PSM and gradually decrease toward the anterior PSM. As cells “move” or are positioned closer to the anterior, they experience decreasing levels of FGF and Wnt, leading to the downregulation of Tbx6. This shift in signaling cues is essential for transitioning cells out of the Tbx6-expressing, unsegmented state, allowing them to prepare for segmentation and somite boundary formation.
- Indirect Activation: While FGF and Wnt signaling are not transcription factors that bind directly to the Tbx6 promoter, they activate Tbx6 expression through downstream effectors and pathways within the cell. These downstream factors likely include various transcriptional regulators that respond to FGF and Wnt signals, which in turn promote Tbx6 transcription in the posterior PSM.
In summary, FGF and Wnt signaling indirectly activate and maintain Tbx6 expression in the posterior PSM, playing an essential role in somitogenesis by supporting Tbx6‘s role in maintaining the unsegmented, mesenchymal state of PSM cells.
(ChatGPT)
In somitogenesis, Tbx6 is a key transcription factor that plays a crucial role in maintaining the undifferentiated, mesenchymal state of cells in the presomitic mesoderm (PSM) and promoting their commitment to the paraxial mesoderm lineage. This role is essential for the orderly formation of somites along the anterior-posterior axis. Here’s a detailed look at Tbx6’s role in somitogenesis:
- Specifying Paraxial Mesoderm Lineage: Tbx6 helps specify cells in the PSM as paraxial mesoderm, which gives rise to somites. In the absence of Tbx6, PSM cells may adopt alternative fates, such as forming neural tissue instead of somites. This lineage specification function is critical in ensuring that the correct tissue types emerge in the right positions during development.
- Maintaining Cells in an Undifferentiated State: Tbx6 expression is high in the posterior PSM, where it keeps cells in an undifferentiated and mesenchymal state, preventing them from undergoing premature segmentation. This maintenance of an undifferentiated state is essential for the proper timing of somite formation. When cells move toward the anterior PSM (注）, Tbx6 expression decreases, which allows them to transition toward epithelialization and segmentation.
- Coordination with FGF and Wnt Signaling: Tbx6 expression is influenced by FGF and Wnt signaling gradients, which are high in the posterior PSM and gradually decrease toward the anterior. These signaling pathways help maintain Tbx6 expression in the posterior PSM, where cells are held in a proliferative, unsegmented state. As cells approach the anterior, decreasing FGF and Wnt signals lead to downregulation of Tbx6, allowing cells to exit the proliferative state and begin the process of segmentation.
- Regulation of Somite Boundary Formation: Although Tbx6 itself does not directly trigger the mesenchymal-epithelial transition (MET) needed for somite boundary formation, it creates the conditions necessary for cells to respond to other cues, such as Mesp2, that initiate boundary formation. As cells exit the Tbx6-expressing region, they become competent to form boundaries and undergo MET, marking the anterior edge of a new somite.
- Interaction with Mesp2 for Rostrocaudal Polarity: Tbx6 indirectly influences the establishment of rostrocaudal (head-to-tail) polarity within somites by interacting with pathways that activate Mesp2 expression in the anterior of each forming somite. Mesp2 expression is critical for defining the anterior-posterior polarity, which is essential for the correct alignment and differentiation of somites. Proper timing of Tbx6 downregulation is necessary for Mesp2 to function appropriately in defining this polarity.
In summary, Tbx6 is essential for maintaining the mesenchymal state and paraxial mesoderm identity of cells in the PSM, coordinating with signaling pathways to control the timing and positioning of somite formation. By controlling when and where cells become competent to segment, Tbx6 ensures the sequential and ordered formation of somites, a fundamental aspect of vertebrate body segmentation.
(ChatGPT)
（注）上の説明でmoveという言葉が使われていたので、てっきり細胞移動が起こると説明しているのかと思い、？？でしたが、よくよく聞いてみたら物理的な移動を意味するmoveではないとのことでした。
In this context, “move toward the anterior PSM” does not imply physical movement of individual cells. Instead, it refers to the positional progression of cells along the anterior-posterior axis within the presomitic mesoderm (PSM) due to developmental processes.
More accurately, what happens is this:
- Cells in the PSM are gradually positioned closer to the anterior as new cells are added posteriorly due to tail bud growth and axial elongation. This process creates a progressive shift without requiring individual cell migration.
- As cells reach the anterior PSM (or the “determination front”), their exposure to signaling gradients changes (with decreased FGF/Wnt and increased RA), which influences gene expression patterns, such as the downregulation of Tbx6.
Thus, a clearer phrasing might be: “As cells are positioned closer to the anterior PSM…”
(ChatGPT)
英語のmoveという言葉には、物理的な移動だけでなく状況の変化を意味する用法もあるというわけです。
In English, the word “move” can indeed have both literal and figurative meanings, depending on the context:
- Literal Meaning (Physical Movement): “Move” often implies physical movement of an object from one place to another. For example, “The car moved down the street” means the car physically traveled along the road.
- Figurative or Conceptual Meaning (Change Without Physical Movement): “Move” can also describe changes that are not physical. For instance:
- Position in a sequence or process: “The project moved to the next phase” implies progression in steps rather than physical movement.
- Change in state or condition: “He moved to a new job role” indicates a change in status or position without physical relocation.
- Shift in time or focus: “We moved the deadline” or “The story moves to a new topic” means a change in timing or subject matter, not a physical shift.
In scientific or technical language, “move” is often used figuratively to describe progression, transition, or reconfiguration within a defined space or system, even if no physical movement occurs. For instance, in biology, when we say cells “move toward differentiation,” it may mean they progress through developmental stages without physically migrating.
(ChatGPT)
以下,ChatGPT 4oの回答ですが、RAはMesp2遺伝子の発現を直接、活性化するわけではないようです。FGFが抑制していたものを解き放つという、間接的な形での作用だとのこと。
In somitogenesis, retinoic acid (RA) plays a key role in setting up the anterior-posterior (A-P) gradient within the presomitic mesoderm (PSM). This gradient interacts with other signaling gradients, particularly those of fibroblast growth factor (FGF) and Wnt, to establish a regulatory environment that is crucial for segmental patterning.
RA does not directly activate the expression of Mesp2, a transcription factor essential for somite boundary formation and rostrocaudal polarity. Instead, RA indirectly influences Mesp2 expression by opposing the posteriorizing effects of FGF and Wnt signaling in the PSM. High levels of FGF and Wnt signaling in the posterior PSM inhibit Mesp2 expression, whereas RA, which is synthesized in the anterior region, helps to repress FGF and Wnt as cells progress anteriorly. This repression allows the anterior PSM cells to express Mesp2 in a specific region, known as the “determination front,” where conditions are favorable for somite segmentation.
Thus, RA’s role in somitogenesis is to help balance the signaling environment in the PSM by counteracting FGF and Wnt, creating a permissive environment for Mesp2 expression at the right place and time, rather than acting as a direct activator of Mesp2.
レチノイン酸とFGF/Wntは拮抗した作用をもつわけですが、具体的に何を抑制するのでしょうか。FGFやWntの遺伝子発現そのものを抑制するわけではなく、例えばFGFシグナリングの要素であるマップキナーゼを脱リン酸化する酵素の遺伝子発現を促進することでFGFシグナリングを弱めたりするのだそうです。またFGF受容体の遺伝子発現を抑制するという報告もあるそう（以下のChatGPTの説明を参照）。
Retinoic acid (RA) opposes FGF and Wnt signaling in the presomitic mesoderm (PSM) through several mechanisms, but it does not primarily act by directly suppressing the transcription of FGF or Wnt genes. Instead, RA counteracts FGF/Wnt signaling by influencing downstream signaling pathways and components in the following ways:
- Inhibition of FGF and Wnt Signaling Pathways:
- RA can indirectly interfere with FGF and Wnt signaling by modulating the expression of downstream targets or components within these pathways.
- For example, RA has been shown to induce the expression of MAP kinase phosphatases (such as Dusp6), which inactivate components of the FGF signaling cascade, thereby reducing FGF signaling activity. This reduces the phosphorylation and activity of key FGF signaling molecules, effectively dampening FGF’s influence in the anterior PSM.
- Antagonistic Gradient Formation:
- RA and FGF/Wnt establish opposing gradients within the PSM. RA is synthesized in the anterior PSM and diffuses toward the posterior, whereas FGF and Wnt are expressed more strongly in the posterior and decrease toward the anterior.
- This spatial gradient positioning allows RA to gradually counterbalance the effects of FGF and Wnt signaling as cells move anteriorly. Rather than directly suppressing FGF or Wnt gene transcription, RA establishes a microenvironment in the anterior PSM where FGF/Wnt signaling becomes progressively less dominant.
- Indirect Regulation of Oscillatory and Segmental Gene Expression:
- RA promotes the expression of genes required for segmentation and differentiation, such as Mesp2, which is necessary for somite boundary formation. This indirectly opposes the role of FGF and Wnt in maintaining cells in an oscillatory, undifferentiated state.
- By promoting segmental gene expression, RA effectively counteracts FGF/Wnt-driven oscillatory gene activity, allowing cells to exit the oscillatory phase as they approach the determination front.
- Suppression of FGF Receptor Expression:
- In some studies, RA has been shown to downregulate the expression of FGF receptors (e.g., Fgfr1), which decreases the responsiveness of cells to FGF signals as they move toward the anterior PSM. This reduction in receptor expression decreases FGF pathway activity without directly impacting FGF ligand expression.
- Regulation of Cell Cycle and Differentiation States:
- RA promotes cell cycle exit and differentiation, which opposes the proliferative effects of FGF and Wnt signaling. By pushing cells toward a differentiated, segmentation-competent state in the anterior PSM, RA helps counteract the posteriorizing effects of FGF and Wnt, which maintain cells in a proliferative, mesenchymal, and unsegmented state.
Summary
In summary, RA opposes FGF and Wnt signaling mainly through indirect mechanisms rather than directly suppressing FGF or Wnt gene transcription. It modulates the signaling environment by affecting downstream signaling components, receptor expression, and promoting differentiation. These actions ensure that the determination front is established at the correct position within the PSM, allowing for the orderly transition from oscillatory to segmented states necessary for somite formation.
(ChatGPT 4o)
In somitogenesis, FGF (Fibroblast Growth Factor) and Wnt signaling interact with Notch signaling to regulate the timing, positioning, and progression of somite formation. Here’s how FGF and Wnt influence Notch signaling within this developmental process:
1. Establishing the Posterior-to-Anterior Gradient
- FGF and Wnt form a gradient across the presomitic mesoderm (PSM), with high concentrations in the posterior PSM (closer to the tail end) and decreasing toward the anterior PSM.
- This gradient creates a “wavefront” that determines where cells are kept in an undifferentiated, oscillatory state versus where they are able to stop oscillating and begin somite formation.
2. Maintaining the Oscillatory State of Notch Signaling in the Posterior PSM
- High FGF and Wnt levels in the posterior PSM promote and maintain oscillatory gene expression, including that of Lfng(lunatic fringe), Hes7, and Delta in the Notch pathway.
- These oscillations are crucial for the “segmentation clock” that drives periodic Notch signaling activity. FGF and Wnt signaling help sustain this clock in the posterior region by keeping cells in a proliferative, undifferentiated state.
3. Influencing the Determination Front
- As cells progress anteriorly in the PSM, they encounter decreasing levels of FGF and Wnt and increasing levels of retinoic acid (RA).
- At a certain threshold—called the determination front—the low levels of FGF and Wnt allow cells to exit the oscillatory Notch signaling state and stabilize their gene expression patterns, leading to somite boundary formation.
- This interaction sets the position at which the oscillatory clock stops, allowing Notch signaling to stabilize and initiate differentiation.
4. Regulating Somite Boundary Formation
- Notch signaling is essential for establishing the boundaries of somites, and the cessation of oscillatory Notch activity is key to determining where boundaries form.
- By controlling the oscillatory vs. stabilized states of Notch signaling, FGF and Wnt ensure that somite boundaries are positioned correctly along the anterior-posterior axis.
5. Temporal and Spatial Coordination of Somite Formation
- Wnt signaling also works alongside FGF to influence the periodicity of the segmentation clock by modulating the expression of clock genes like Hes7 and Lfng.
- By maintaining cells in an oscillatory state until they reach the determination front, FGF and Wnt ensure that Notch signaling follows a precise temporal pattern that matches the physical positioning along the body axis, contributing to the regular, sequential formation of somites.
Summary
In somitogenesis, FGF and Wnt signaling maintain Notch signaling in an oscillatory state in the posterior PSM, essential for the segmentation clock. As cells move toward the anterior, lower FGF/Wnt levels allow them to exit this oscillatory state at the determination front, where Notch signaling stabilizes to form somite boundaries. This regulation ensures precise timing and spatial organization in somite formation.
(ChatGPT)
下の論文のまとめの図がわかりやすいと思います。
12 July 2011 The Wnt3a/β-catenin target gene Mesogenin1 controls the segmentation clock by activating a Notch signalling program Nature Communications volume 2, Article number: 390 (2011) https://www.nature.com/articles/ncomms1381
周期的に変動する遺伝子がいくつかありますが、どれが一番の発信源といえるのでしょうか。Deltの遺伝子発現には周期性がありますが、Notchにはないようです。ノッチシグナリングとしては周期性があるという言い方は可能です。
下の論文の図をみるとノッチの細胞内ドメインNICDは変動をしているようです。
Fgf4 maintains Hes7 levels critical for normal somite segmentation clock function Nov 19, 2020 https://doi.org/10.7554/eLife.55608 https://elifesciences.org/articles/55608
The vertebrate Embryo Clock: Common players dancing to a different beat Front. Cell Dev. Biol., 11 August 2022 Sec. Morphogenesis and Patterning Volume 10 – 2022 | https://doi.org/10.3389/fcell.2022.944016
In the oscillatory system of somitogenesis, Notch signaling is generally considered the most upstream component that initiates and regulates the segmentation clock, of which HES genes (like Hes1 and Hes7) are key downstream oscillatory elements.
Reasons Why Notch is Upstream of HES
- Activation of HES Genes by Notch Signaling:
- Notch signaling is activated when Delta ligands bind to Notch receptors on neighboring cells. This interaction triggers the Notch intracellular domain (NICD) to be released and translocate to the nucleus, where it activates the transcription of downstream target genes, including the HES family (e.g., Hes1, Hes7).
- Thus, Notch signaling initiates the oscillatory expression of HES genes in the presomitic mesoderm (PSM).
- Synchronization Role of Notch:
- Notch signaling is also critical for synchronizing oscillations between neighboring cells in the PSM, ensuring coordinated oscillatory gene expression across cells that will form a somite boundary. This intercellular communication mediated by Notch and Delta helps maintain the rhythmic, synchronized oscillations of HES genes across the tissue.
- Regulation by FGF and Wnt Gradients:
- Notch activity, influenced by upstream FGF and Wnt gradients, helps establish the oscillatory environment in the posterior PSM. These gradients set up the context for Notch signaling to initiate the oscillations, positioning Notch as a primary regulator in the segmentation clock.
- Feedback Mechanism with HES Genes:
- HES proteins, once expressed, participate in a negative feedback loop to repress their own transcription and the transcription of other oscillatory genes. However, without initial Notch signaling, this feedback loop in HES genes would not be activated, making Notch the upstream trigger.
Summary
In somitogenesis, Notch signaling is the most upstream component of the oscillatory system, activating and coordinating the expression of HES genes and other oscillatory genes within the segmentation clock. HES genes then function in feedback loops as downstream effectors, creating the cyclic gene expression patterns essential for somite formation.
Notchは周期的に変動しないことに関して
In somitogenesis, Notch gene expression itself does not oscillate like Delta; rather, Notch signaling activity oscillates due to the oscillatory expression of its ligands (like Delta) and downstream effectors (such as HES genes).
Here’s how Notch signaling oscillations occur without Notch gene expression itself being oscillatory:
- Constant Notch Receptor Expression:
- The expression of Notch receptors (such as Notch1 in mice) remains relatively stable in the presomitic mesoderm (PSM) and does not oscillate. Instead, the Notch receptor is always present on the cell surface, ready to respond to Delta ligand binding.
- Oscillatory Delta Ligand Expression:
- Delta ligands (e.g., Dll1 in mice) do oscillate in their expression, driven by the segmentation clock. When Delta levels rise on the surface of one cell, it binds to Notch receptors on neighboring cells, activating Notch signaling in those cells in an oscillatory manner.
- This oscillatory expression of Delta ligand initiates periodic activation of Notch signaling in adjacent cells, creating a synchronized oscillation of Notch pathway activity across the PSM.
- Oscillatory Downstream Targets (HES Genes):
- The activation of Notch by oscillating Delta triggers the expression of downstream targets like HES genes (e.g., Hes1, Hes7), which also oscillate. HES proteins provide negative feedback that helps control the oscillatory cycle of Delta and other segmentation clock genes.
- Intercellular Synchronization:
- The stable presence of Notch receptors and the oscillatory expression of Delta enable the synchronized activation of Notch signaling across neighboring cells. This intercellular communication is critical for maintaining coordinated oscillations in the PSM, which ultimately determine the timing and location of somite boundaries.
Summary
While Notch gene expression itself does not oscillate in somitogenesis, the Notch signaling pathway oscillates due to the periodic expression of Delta ligands and HES gene feedback loops. This oscillatory signaling is essential for the segmentation clock and synchronized somite formation across the presomitic mesoderm.
Notch Signaling Pathway | Purpose and Mechanism JJ Medicine チャンネル登録者数 107万人
デルタは遺伝子発現の変動が知られています。
Oscillatory control of Delta-like1 in cell interactions regulates dynamic gene expression and tissue morphogenesis Genes & Dev. 2016. 30: 102-116 https://genesdev.cshlp.org/content/30/1/102/F1.expansion.html
Delta-Notchシグナリングは、周期的な遺伝子発現変化を生じさせることに必要なわけではないそうです。デルタノッチシグナリングを阻害しても、個々の細胞は遺伝子の周期的な発現を示すそう。ただし、それらの細胞の間での「同期」が壊れてしまうのだそうです。つまり、Delta-notchの役割は、細胞集団間での振動の同期ということのようです。
10 May 2024 Oscillatory control of embryonic development Development (2024) 151 (9): dev202191. https://journals.biologists.com/dev/article/151/9/dev202191/347230/Oscillatory-control-of-embryonic-development
下の動画はノッチシグナリングレポーターによるノッチシグナリングの可視化。レポーターの設計の原理は下の論文を参照。
Notch intracellular domain (NICD), which translocates to the nucleus and associates with the DNA binding protein CSL [CBF1/RBPjκ/Su(H)/Lag1] to activate gene expression. luciferase complementation imaging (LCI) reporters for NICD-CSL association https://pubmed.ncbi.nlm.nih.gov/21775282/
Oscillatory Notch signalling during somite boundary formation, imaged using SPIM The Company of Biologists チャンネル登録者数 5930人
同様にChatGPTの回答です。
In somitogenesis, the Notch-Delta signaling pathway is essential for coordinating the timing and spatial organization of somite formation. This pathway functions as part of a molecular “segmentation clock,” which ensures that somites form in a regular, sequential manner along the anterior-posterior (A-P) axis of the developing embryo.
Key roles of Notch-Delta signaling in somitogenesis include:
- Oscillatory Gene Expression: Notch-Delta signaling drives oscillations of specific “clock genes” in the presomitic mesoderm (PSM), such as Hairy1, Hes7, and others, which cycle in a periodic manner. This oscillation occurs with a set frequency, creating a wavefront of gene expression that moves through the PSM. This “segmentation clock” is crucial for determining the periodicity of somite formation.
- Synchronization of Cells: Notch-Delta signaling helps synchronize the oscillatory behavior of neighboring cells in the PSM. Through intercellular communication, Delta ligands from one cell bind to Notch receptors on adjacent cells, coordinating their segmentation clocks. This synchronization ensures that cells within a future somite boundary express genes likeMesp2 together, enabling the precise formation of somites.
- Boundary Formation and Rostrocaudal Polarity: Notch signaling is critical in specifying somite boundaries and establishing rostrocaudal (head-to-tail) polarity within each somite. At the determination front, where somites bud off from the PSM, Notch signaling interacts with factors like Mesp2 to trigger the epithelialization process that creates the somite boundary. The periodic activation of Notch also helps establish differential gene expression within each somite, giving them a distinct anterior and posterior polarity.
- Coordination with FGF and RA Gradients: Notch-Delta signaling is integrated with FGF and RA gradients in the PSM, which help establish the anterior-posterior axis. Notch oscillations occur within a field shaped by these gradients, defining the region where somites form (the “determination front”). As the PSM cells move out of the FGF-rich posterior region toward the RA-rich anterior region, Notch signaling cues become crucial in determining where and when a somite boundary will form.
In summary, Notch-Delta signaling coordinates the temporal and spatial patterning of somite formation by driving oscillatory gene expression, synchronizing cell behaviors, defining boundaries, and interacting with other signaling pathways. This fine-tuned regulation is critical for forming the segmented structure of the vertebrate body.
(ChatGPT)
In somitogenesis, Delta (a ligand in the Notch signaling pathway) exhibits oscillatory gene expression primarily through its regulation by the segmentation clock, which is driven by negative feedback loops involving oscillatory genes like Hes family members (e.g., Hes1 and Hes7). Here’s a breakdown of the mechanism behind Delta’s oscillatory expression:
- Notch-Delta Signaling and Feedback Loop:
- Delta ligands (e.g., Dll1 in mice) are part of the Notch signaling pathway, which is crucial for coordinating the segmentation clock across neighboring cells in the presomitic mesoderm (PSM).
- When Delta binds to Notch receptors on neighboring cells, it activates the Notch pathway, leading to the expression of downstream genes, including Hes genes. This interaction synchronizes oscillatory gene expression between neighboring cells, which is essential for organized somite formation.
- Hes Genes as Transcriptional Repressors:
- Hes genes, activated by Notch signaling, function as transcriptional repressors and play a key role in generating oscillations. For example, Hes7 represses its own transcription as well as that of Delta by binding to their promoters, creating a negative feedback loop.
- After Hes proteins are synthesized, they inhibit further transcription of their own gene and Delta. As Hes proteins degrade, repression is lifted, allowing Delta and Hes transcription to restart, resulting in a cycle of oscillatory expression.
- Negative Feedback with a Time Delay:
- This oscillatory behavior relies on a time delay in the negative feedback loop, which is a function of mRNA and protein synthesis, degradation rates, and cellular delays in transcriptional repression.
- The degradation of Hes proteins and mRNA is relatively rapid, and once they are sufficiently degraded, repression is released, allowing a new cycle of Delta and Hes expression to begin. This delay-dependent negative feedback is critical for generating stable oscillations.
- Synchronization Across Cells via Notch-Delta Interactions:
- Notch-Delta signaling between neighboring cells helps synchronize their oscillatory patterns. Delta expression in one cell activates Notch in an adjacent cell, which in turn influences Hes gene expression in that neighboring cell, creating coordinated oscillations.
- This intercellular coupling ensures that Delta expression oscillates synchronously across cells in the PSM, which is essential for creating uniform, periodic somite boundaries.
- Interaction with FGF and Wnt Gradients:
- Delta oscillations also interact with the FGF and Wnt signaling gradients across the PSM. These gradients are high in the posterior PSM and decrease toward the anterior, modulating the pace and location of Delta oscillations.
- As cells move toward the anterior PSM, where FGF and Wnt levels are lower, Delta oscillations slow and eventually stabilize, marking the transition point where somite boundaries begin to form.
Summary
Delta’s oscillatory expression in somitogenesis is driven by a negative feedback loop involving Hes genes, with a time delay that produces cyclical repression and derepression. This oscillation is synchronized across cells by Notch-Delta signaling between neighboring cells, and modulated by the FGF and Wnt gradients to ensure coordinated somite formation.
(ChatGPT)
lunatic fringe遺伝子はNOTCHシグナリングによって直接、活性化されるようです。lunatic fringeの働きは抑制的で、NOTCHを抑制します。
Lunatic fringe (Lfng) gene expression oscillates in somitogenesis, and it is driven by Notch signaling. Here’s a detailed look at why and how this oscillation occurs:
1. Role of Notch Signaling in Driving Lunatic Fringe Oscillations
- Notch Activation: The oscillatory expression of Lfng is a result of periodic Notch signaling activation in the presomitic mesoderm (PSM). Notch signaling is activated by ligands such as Delta-like 1 (Dll1), which bind to Notch receptors on neighboring cells.
- Transcriptional Activation of Lfng: When Notch receptors are activated, they release the Notch intracellular domain (NICD), which translocates to the nucleus and activates transcription of target genes, including Lfng.
- Negative Feedback and Oscillations: Lfng itself plays a role in modulating Notch signaling by glycosylating the Notch receptor, which modifies its responsiveness to Delta ligands. This feedback can suppress further activation of Notch in a periodic manner. This creates a negative feedback loop where Lfng expression periodically increases and decreases, creating oscillations.
2. Why Lunatic Fringe Needs to Oscillate
- Coordination with the Segmentation Clock: Oscillatory expression of Lfng is synchronized with other segmentation clock genes (such as Hes7), ensuring that Notch signaling oscillates in a coordinated manner across the PSM. This synchronization is critical for the precise timing of somite boundary formation.
- Control of Somite Boundary Formation: Lfng oscillations help regulate when and where Notch signaling is active along the anterior-posterior axis, contributing to the orderly segmentation needed for somite boundary formation.
3. Interaction with Other Signals
- Wnt and FGF Gradients: While Notch signaling directly drives Lfng oscillations, the activity of Notch signaling itself is modulated by Wnt and FGF gradients across the PSM. High levels of Wnt and FGF in the posterior PSM sustain the oscillatory environment, maintaining Notch signaling activity. As cells move toward the anterior and encounter lower levels of Wnt and FGF, they begin to stabilize their gene expression patterns, including Lfng, as they prepare for somite formation.
- Negative Feedback in the Notch Pathway: In addition to Lfng, other Notch pathway components like Hes genes also oscillate and contribute to the timing and coordination of Notch-driven processes.
Summary
Notch signaling drives the oscillatory expression of Lunatic fringe (Lfng) in somitogenesis. The oscillations in Lfng are a result of Notch activation by ligands like Delta, combined with a negative feedback loop where Lfng itself modulates Notch receptor sensitivity. This oscillation is synchronized with other segmentation clock genes, allowing precise timing and spatial organization for somite boundary formation. The stability of Lfng oscillations depends on Notch signaling, which is further modulated by posterior Wnt and FGF gradients.
(ChatGTP 4o)
Lunatic fringe (often abbreviated as Lfng) is a gene that encodes an enzyme known as a glycosyltransferase, which modifies the Notch receptor, influencing how it responds to Delta and other Notch ligands. It plays an important role in regulating Notch signaling oscillations during somitogenesis. Here’s a breakdown of lunatic fringe’s function, its name, and how it fits into the Notch signaling pathway.
1. What Lunatic Fringe Does
- Role in Notch Signaling: Lunatic fringe modifies the Notch receptor by adding sugars to it (glycosylation). This modification changes the sensitivity of Notch to its ligands, including Delta-like (Dll1) and Jagged ligands.
- Oscillatory Expression: Like Delta and HES genes, Lfng expression oscillates in the presomitic mesoderm (PSM) as part of the segmentation clock. These oscillations help regulate the timing of Notch activation, contributing to the rhythmic formation of somites.
- Function in Somite Formation: By modulating Notch signaling, lunatic fringe helps maintain the periodic activation of Notch in a way that coordinates somite segmentation across neighboring cells.
2. Literal Meaning of the Name “Lunatic Fringe”
- The name “lunatic fringe” is somewhat playful, inspired by the idea of an “edge” or “fringe” that behaves in a periodic or “lunatic” (crazy, unpredictable) manner.
- Fringe refers to the family of fringe proteins (other members include manic fringe and radical fringe), which all influence Notch signaling by modifying the Notch receptor’s response to its ligands.
- Lunatic implies something erratic or periodic, likely a nod to the cyclic or “oscillatory” behavior seen in Notch signaling and segmentation, somewhat like the moon’s periodic phases (the root “luna” refers to the moon).
Summary
Lunatic fringe is a glycosyltransferase enzyme that modifies the Notch receptor, affecting how it responds to ligands like Delta. Its oscillatory expression is crucial for somite formation in somitogenesis. While it is neither Delta nor HES, it plays a critical role in controlling the oscillations of Notch signaling that drive the segmentation clock. The name “lunatic fringe” reflects the periodic or “fringe” nature of its action in the oscillatory system of somitogenesis.
(ChatGPT)
下の動画はマウス胚で体節が形成されるごとに遺伝子発現が変動するLunatic fringeという遺伝子。
Imaging Clock Oscillations in the Mouse Embryo Cell Press チャンネル登録者数 2.58万人
Wave of gene expression in an embryo European Molecular Biology Laboratory (EMBL) チャンネル登録者数 1.34万人
HES1はHairy and Enhancer of Split 1の略で、Hairy遺伝子ファミリーの一部です。 HES1は、哺乳類のHairyホモログであり、Notchシグナル伝達経路によって誘導される転写因子です。もともとHairyはショウジョウバエ（Drosophila）で発見され、発生中の体節形成や遺伝子発現の制御に関与しています。哺乳類では、HES1やHES7などが体節形成や神経発生の際に遺伝子発現の振動を担い、体の発生において重要な役割を果たしています。 このため、HES1は「哺乳類のHairy遺伝子の一種」として位置づけられ、振動的な発現パターンを示す遺伝子として、発生学的な研究で頻繁に参照される遺伝子の1つです。
Avian hairy Gene Expression Identifies a Molecular Clock Linked to Vertebrate Segmentation and Somitogenesis Isabel Palmeirim1 ∙ Domingos Henrique2,§ ∙ David Ish-Horowicz2 ∙ Olivier Pourquié CELL Volume 91, Issue 5p639-648November 28, 1997 https://www.cell.com/fulltext/S0092-8674%2800%2980451-1
HESは転写抑制因子として働きますが、標的遺伝子の中には自分自身も含まれています。HESは分解速度が速い（20分程度）ため、タンパク質が分解されてなくなれば、転写抑制の抑制が解けてまたタンパク質が翻訳されます。その繰り返しによって、周期的な変動が生じるというわけです。抑制因子が「時間遅れ」をもって現れるというのがミソです。また抑制因子の分解が速いというのも大事。この論文、
Autoinhibition with Transcriptional Delay: A Simple Mechanism for the Zebrafish Somitogenesis Oscillator Julian Lewis Current Biology Volume 13, Issue 16, 19 August 2003, Pages 1398-1408 Journal home page for Current Biology Article https://doi.org/10.1016/S0960-9822(03)00534-7 https://www.sciencedirect.com/science/article/pii/S0960982203005347
にシミュレーションの結果や微分方程式などが掲載されていました。
https://www.sciencedirect.com/science/article/pii/S0960982203005347
サプリメンターリーデータにタンパク質分解速度などのパラメータが全部乗っている（マセマティカのコードが掲載）ので、自分でPythonなどで再現できます。ChatGPTに頼んだら動くコードを秒で書いてくれました。
ChatGPTにまとめてもらいます。
In somitogenesis, Notch, WNT, and FGF signaling pathways all play critical roles, but their oscillatory behaviors vary:
1. Notch Pathway
- Oscillatory Behavior: YES
- Genes in the Notch signaling pathway, such as Hes/Hairy family transcription factors, exhibit robust oscillatory expression in the presomitic mesoderm (PSM).
- Oscillations are driven by transcriptional feedback loops:
- Hes1 represses its own transcription, leading to periodic expression cycles.
- These oscillations are synchronized among neighboring cells through Notch-mediated intercellular communication.
2. WNT Pathway
- Oscillatory Behavior: YES
- Components of the WNT signaling pathway, like Axin2, exhibit oscillatory expression.
- Oscillations in the WNT pathway are driven by negative feedback loops involving β-catenin stabilization and Axin2 expression.
- These oscillations are slower compared to Notch oscillations and are thought to interact with both Notch and FGF pathways.
3. FGF Pathway
- Oscillatory Behavior: CONTROVERSIAL / INDIRECT
- FGF signaling itself is not directly oscillatory, but its downstream targets, such as Dusp6 and Spry4, show oscillatory expression in some species (e.g., zebrafish).
- FGF’s gradient along the PSM establishes a “wavefront” rather than an intrinsic oscillator. However, this wavefront interacts with the oscillatory clock (Notch and WNT) to define the periodic segmentation of somites.
Summary of Oscillatory Behavior
Pathway Oscillatory Expression Key Oscillatory Genes Notch Yes Hes1, Hes7, Lfng WNT Yes Axin2, Dkk1 FGF Indirect/Controversial Dusp6, Spry4 (in zebrafish and some species)
Interaction Between Pathways
- Coupling of Oscillators:
- Notch, WNT, and (indirectly) FGF oscillators are coupled to ensure coordinated somite formation.
- Wavefront Mechanism:
- The FGF signaling gradient works as a “wavefront” that interacts with the Notch and WNT “clock” to regulate the timing and position of somite segmentation.
In conclusion, Notch and WNT pathways show clear oscillatory gene expression, while FGF signaling oscillations are species-dependent and often secondary to other oscillatory systems. Together, these pathways synchronize to drive the somitogenesis clock.
発現レベルが振動する遺伝子の遺伝子産物とモルフォゲンとして働く遺伝子産物はどのように相互作用するのでしょうか。ChatGPTに訊きました。
The FGF signaling gradient interacts with the oscillatory clock genes (primarily from the Notch and WNT pathways) to coordinate the segmentation process during somitogenesis. Here’s a detailed explanation of this interaction:
1. Overview of the FGF Gradient
- Gradient Formation:
- The FGF signaling pathway establishes a posterior-to-anterior gradient in the presomitic mesoderm (PSM).
- FGF8 (and to some extent FGF4) is produced in the posterior PSM and diffuses anteriorly, creating a concentration gradient.
- Degradation of FGF mRNA (e.g., by retinoic acid signaling from the anterior) ensures a sharp decline in FGF signaling towards the anterior.
- Function:
- The FGF gradient defines a wavefront, also known as the “determination front,” which interacts with the oscillatory clock genes to determine the spatial boundary for somite formation.
2. Oscillatory Clock Genes
- Genes from the Notch (e.g., Hes7) and WNT (e.g., Axin2) pathways oscillate periodically in the PSM.
- These oscillations are synchronized within and between cells, creating waves of gene expression that travel posterior-to-anterior in the PSM.
3. Interaction Between FGF Gradient and Clock Genes
The FGF gradient interacts with the clock genes in a spatiotemporal manner to regulate somite segmentation:
(1) Threshold Mechanism
- Anterior FGF Threshold:
- Cells in the PSM “read” the FGF gradient. When the FGF concentration drops below a certain threshold, oscillatory clock gene expression stops, and the cells are primed to form a somite.
- This threshold defines the anterior boundary of a new somite.
- Coupling with Oscillations:
- The clock genes, which oscillate in a periodic manner, are interpreted by the wavefront (set by the FGF gradient) to determine when and where segmentation occurs.
(2) Synchronization of Timing
- The oscillatory cycles of clock genes determine the timing of somite formation.
- The FGF gradient sets the spatial position where the next somite boundary will form.
(3) WNT-FGF Feedback Loop
- WNT-Dependent Regulation of FGF:
- WNT signaling in the posterior PSM maintains FGF expression. This ensures the FGF gradient remains sharp and dynamic.
- FGF’s Effect on WNT:
- FGF signaling can modulate the WNT pathway through feedback loops, ensuring that WNT-driven oscillations are synchronized with the FGF gradient.
4. Molecular Interactions
- FGF and Clock Gene Regulation:
- FGF Downstream Genes:
- Dusp6, Spry2, and Spry4 are feedback regulators of FGF signaling. These genes oscillate in the posterior PSM in some species (e.g., zebrafish), indirectly linking FGF signaling to the oscillatory clock.
- Clock Gene Impact on FGF Gradient:
- The segmentation clock genes regulate the Mesp2 expression at the wavefront, which is critical for determining somite boundaries. Mesp2 expression is tightly regulated by the interplay between the FGF gradient and Notch signaling.
5. Spatiotemporal Dynamics
The interaction of the FGF gradient and the oscillatory clock genes ensures that:
- Timing:
- The oscillatory clock sets the periodicity of somite formation (e.g., ~90 minutes in mice).
- Position:
- The FGF gradient sets the spatial location where somite boundaries form.
- Coordination:
- As the FGF gradient moves posteriorly due to axial elongation, it provides a moving reference for the oscillations to interact with.
6. Evolutionary and Experimental Insights
- Species-Specific Differences:
- In zebrafish, FGF signaling shows oscillatory expression (e.g., Dusp6), whereas in mice, FGF is primarily a static gradient with a wavefront function.
- Perturbation Studies:
- Disrupting FGF signaling (e.g., by inhibiting FGF receptors) abolishes the ability of the wavefront to interact with oscillatory clock genes, leading to defective somite formation.
7. Summary
- The FGF gradient serves as a wavefront that interacts with oscillatory clock genes to spatially and temporally coordinate somite formation.
- Key mechanisms:
- FGF concentration below a threshold stops oscillations and triggers somite segmentation.
- The interplay between FGF and WNT signaling ensures proper maintenance of the gradient and its alignment with clock oscillations.
- This dynamic interaction ensures the periodic formation of somites, which is critical for proper vertebrate body segmentation.
This interaction showcases a remarkable example of how temporal oscillations and spatial gradients integrate to create precise developmental patterns.
FGFの濃度が下がるとなぜ振動が止まるのでしょうか。これもChatGPTに訊いてみます。
The drop in FGF concentration triggers the cessation of oscillations in the presomitic mesoderm (PSM) because the FGF signaling pathway regulates the transcriptional and translational dynamics necessary for maintaining the oscillatory clock. The mechanisms underlying this process involve threshold regulation, cross-talk with WNT and Notch pathways, and transcriptional control of critical genes. Below is a detailed explanation:
1. Role of FGF in Maintaining Oscillations
- FGF signaling sustains the expression of oscillatory clock genes (e.g., Hes7 in the Notch pathway and Axin2 in the WNT pathway) in the posterior PSM.
- High FGF concentration in the posterior:
- Promotes active transcription of clock genes.
- Supports the dynamic feedback loops necessary for oscillations.
- As cells move anteriorly in the PSM, they experience progressively lower FGF concentrations due to the FGF gradient.
2. Mechanisms for Oscillation Cessation
(1) Threshold Mechanism
- FGF concentration-dependent transition:
- FGF levels drop below a threshold as cells migrate anteriorly.
- This low FGF signaling environment disrupts the transcriptional feedback loops that drive oscillations.
(2) FGF’s Regulation of Clock Genes
- Notch Pathway Genes:
- FGF signaling supports the periodic expression of Hes7 by regulating upstream Notch pathway components.
- Loss of FGF signaling destabilizes Hes7’s negative feedback loop, halting oscillations.
- WNT Pathway Genes:
- FGF maintains WNT signaling in the posterior PSM, ensuring oscillatory behavior of genes like Axin2.
- A reduction in FGF signaling decreases WNT activity, which is critical for oscillation maintenance.
(3) FGF-Dependent Transcriptional Control
- Mesp2 Activation:
- When FGF concentration drops below the threshold, the transcription factor Mesp2 is upregulated in the anterior PSM.
- Mesp2 represses genes involved in the clock’s oscillatory machinery (e.g., Hes7), promoting stabilization rather than continued oscillation.
- FGF Suppresses Mesp2:
- High FGF levels inhibit Mesp2 expression in the posterior PSM, keeping cells in an oscillatory state.
- Loss of this suppression allows Mesp2 to act, halting oscillations and initiating somite boundary formation.
(4) Cross-Talk with Retinoic Acid (RA)
- FGF and RA gradients are antagonistic:
- Retinoic acid (RA), which is concentrated in the anterior PSM, counteracts FGF signaling.
- As cells move anteriorly and experience lower FGF levels, RA activity increases, further suppressing clock gene oscillations.
- RA promotes Mesp2 expression, solidifying the cessation of oscillations.
(5) Epigenetic and Translational Changes
- FGF signaling maintains the permissive transcriptional and translational environment necessary for oscillations by:
- Regulating the activity of transcription factors like Tbx6, which is essential for maintaining the oscillatory state.
- Supporting ribosomal function and RNA processing for rapid feedback regulation.
- A drop in FGF signaling results in transcriptional repression and slowed feedback dynamics, halting oscillatory behavior.
3. Experimental Evidence
- FGF Inhibition Studies:
- Pharmacological inhibitors of FGF signaling (e.g., SU5402) in mouse and zebrafish embryos disrupt oscillations in the PSM and prematurely stabilize the expression of clock genes.
- FGF-RA Interaction Studies:
- Altering the balance of FGF and RA gradients shifts the location of oscillation cessation, confirming their antagonistic role in regulating the clock.
- Genetic Models:
- Knockout models of FGF pathway components (e.g., Fgf8 or Sprouty genes) show defective segmentation and premature cessation of oscillations.
4. Conceptual Summary
- The FGF gradient functions as a spatial timer:
- In regions of high FGF, oscillations are active due to the maintenance of feedback loops.
- In regions of low FGF, feedback loops are destabilized, leading to oscillation cessation.
- The cessation of oscillations is coupled with:
- Mesp2 activation.
- Transition to a stabilized gene expression state necessary for somite boundary formation.
This mechanism ensures that the oscillatory “clock” and the spatial “wavefront” are tightly coordinated, allowing for precise somite segmentation.
ChatGPTの回答。
The specification of the caudal (posterior) boundary of a newly formed somite is tightly regulated by several interacting signaling pathways and genes. Key signals and regulatory elements involved include:
- Notch Signaling: Notch signaling is crucial for determining the rostrocaudal (head-to-tail) polarity within each somite. In particular, Mesp2, a downstream target of Notch, is expressed in the anterior portion of the forming somite. This differential expression contributes to defining the boundary between the rostral (anterior) and caudal (posterior) compartments of each somite. Notch activity helps restrict Mesp2 expression, thereby influencing the formation of the boundary at the caudal end.
- FGF and Wnt Gradients: Fibroblast Growth Factor (FGF) and Wnt signaling gradients are high in the posterior presomitic mesoderm (PSM) and gradually decrease toward the anterior. As cells move anteriorly within the PSM, they leave the influence of high FGF and Wnt levels, which allows for changes in gene expression critical for somite boundary formation. These gradients work together with Notch signaling to ensure that cells in the anterior PSM can form somites with distinct anterior and posterior regions.
- RA (Retinoic Acid) Gradient: Retinoic acid, produced in the anterior PSM, creates a gradient opposing that of FGF and Wnt. This gradient helps create a permissive environment in the anterior PSM for somite boundary formation. The balance between RA and FGF/Wnt gradients helps set up the “determination front” where somites form, with the anterior boundary of the new somite influenced by RA and the posterior boundary still affected by FGF and Wnt signaling.
- Tbx6 and Ripply Genes: Tbx6, a transcription factor expressed in the posterior PSM, helps maintain the unsegmented state of cells in this region. As cells reach the determination front, Tbx6 expression is downregulated in a process partly mediated by the Ripply genes, which are also regulated by Mesp2. This downregulation allows for the differentiation of the anterior and posterior compartments of the somite, specifying the caudal boundary of the newly formed somite.
- Ephrin/Eph Signaling: Once the somite boundary has been established, Ephrin/Eph signaling becomes active at the newly formed boundary, reinforcing segmentation and boundary stabilization. Ephrin ligands are typically expressed in the posterior cells, while Eph receptors are in the anterior cells of the next forming somite. This interaction helps create a clear caudal boundary for each somite.
Together, these signals and gene interactions establish the caudal boundary of the somite, ensure correct rostrocaudal polarity, and contribute to the distinct segmentation pattern necessary for organized vertebrate body plan development.
下のChatGPTの回答によれば、尾側の境界が先にできて、そのあとで吻側の境界ができるのだそうです。意外でした。自分は逆かと思いました。
When a new somite forms, the establishment of its anterior and posterior boundaries is a successive process rather than a simultaneous one.
Here’s a breakdown of how these boundaries form:
- Posterior Boundary Formation: The posterior boundary of the new somite is defined first. This boundary is established as cells in the presomitic mesoderm (PSM) reach the “determination front,” where they encounter conditions favorable for somite segmentation. At this front, signals such as reduced FGF and Wnt levels (alongside elevated RA levels in the anterior PSM) allow these cells to undergo the segmentation process. Notch signaling also plays a crucial role here, helping to define the initial boundary.
- Anterior Boundary Formation: After the posterior boundary is defined, the anterior boundary of the forming somite is established. This is typically marked by the differential expression of genes like Mesp2, which is initially restricted to the rostral (anterior) half of the somite. This expression leads to the establishment of the anterior boundary, defining the complete somite with distinct anterior and posterior halves.
This stepwise process ensures that each somite forms with the correct orientation and rostrocaudal polarity. By establishing boundaries sequentially, the somitogenesis machinery ensures that each somite is properly aligned with its neighboring segments, creating the precise, segmented structure essential for vertebrate body patterning.
(ChatGPT)
In somitogenesis, the Mesenchymal-Epithelial Transition (MET), which is essential for somite boundary formation, involves the gene Paraxis as well as other key players. Here’s how these genes contribute to the MET process:
- Paraxis: Paraxis is a basic helix-loop-helix (bHLH) transcription factor that is crucial for the epithelialization of somites. Paraxis promotes the transition of mesenchymal cells in the presomitic mesoderm (PSM) to a more epithelial, organized structure within each somite. In Paraxis mutants, somites fail to undergo proper epithelialization, resulting in disorganized, mesenchymal-like structures rather than well-defined somites.
- E-cadherin and N-cadherin: Cadherins are adhesion molecules essential for establishing and maintaining the epithelial structure of cells. During MET, cells in the forming somite upregulate cadherins (especially N-cadherin), promoting cell-cell adhesion and the formation of a segmented epithelial structure.
- Fibronectin and Integrins: These extracellular matrix (ECM) components and receptors help stabilize the epithelial structure within somites. Fibronectin provides structural support in the ECM, and integrins on the cell surface interact with fibronectin to promote cell adhesion and polarity, reinforcing the MET process.
- Notch Signaling and Mesp2: Notch signaling, via Mesp2, also influences the MET process. Mesp2 expression in the anterior portion of each forming somite contributes to boundary formation and epithelialization, working in concert with Paraxis and cadherins to establish the segmented, epithelial state.
Together, these genes and molecular pathways drive the mesenchymal-epithelial transition in somitogenesis, enabling the formation of organized, epithelialized somites essential for the segmented structure of the vertebrate body plan.
(ChatGPT)
上の説明では体節の前半分という話がでてきましたが、後ろ半分に関してはどうなのでしょうか。
In the posterior portion of each forming somite, the mesenchymal-epithelial transition (MET) and establishment of somite boundaries are regulated by specific genes that contribute to the segmentation and polarization of somites. Key genes essential for MET and somite formation in the posterior region include:
- Mesp2: Mesp2 is a crucial transcription factor that plays a role in defining the anterior-posterior polarity within each somite. In the anterior portion of a forming somite, Mesp2 expression helps initiate MET and boundary formation. This, in turn, indirectly defines the posterior boundary by establishing a sharp contrast with the posterior region, which has lower Mesp2 expression.
- Notch Signaling Pathway: The Notch pathway is integral to maintaining the segmentation clock and ensuring that the posterior PSM cells enter a synchronized cycle of differentiation and segmentation. Notch signaling also helps specify the rostrocaudal polarity within each somite. In particular, genes such as Dll1 (Delta-like 1) and Lfng (Lunatic fringe) within the Notch pathway show oscillatory expression patterns that help set the timing and boundary for somite formation. Notch activity is crucial for the posterior portion to remain distinct from the anterior and to respond appropriately during the MET.
- Uncx4.1 (Uncx): Uncx4.1 is expressed specifically in the posterior half of each somite after it forms. This transcription factor helps maintain the identity and structure of the posterior compartment and is essential for establishing the rostrocaudal (head-to-tail) polarity within the somite. It represses anterior markers and maintains the distinct identity of the posterior portion.
- Tbx6: Tbx6, expressed in the posterior PSM, helps keep cells in an undifferentiated state until they reach the determination front, where FGF and Wnt levels are reduced. Once cells are positioned within the forming somite, Tbx6 downregulation is required for epithelialization to proceed in the anterior, which indirectly helps establish the posterior identity in cells that did not fully downregulate Tbx6 initially.
- Ephrin/Eph Signaling: Ephrin ligands (often expressed in the posterior region) and Eph receptors (expressed in the anterior of the next somite) create a boundary of cell-cell repulsion between somites. This signaling enforces a clear distinction between the posterior of one somite and the anterior of the next. Ephrin/Eph interactions are essential for the stabilization and integrity of the posterior boundary.
In summary, genes like Mesp2, Uncx4.1, and components of the Notch, Ephrin/Eph, and FGF signaling pathways collectively ensure that the posterior portion of each somite retains its unique identity and undergoes the MET process in a way that preserves somite polarity and structure. These interactions allow the proper segmentation of the vertebrate body plan during somitogenesis.
(ChatGPT)
The regulation of oscillatory genes like hairy (known as Hes genes in vertebrates, such as Hes7 in mice) in somitogenesis is governed by a network of signaling pathways and genes that collectively form the segmentation clock. This clock drives the periodic expression of oscillatory genes, which is essential for the timing of somite formation. Key regulators of these oscillatory genes include:
- Notch Signaling Pathway: Notch signaling plays a central role in regulating oscillatory gene expression in the presomitic mesoderm (PSM). The activation of Notch receptors by ligands such as Delta-like 1 (Dll1) triggers a cascade that induces the transcription of Hes7, a gene that oscillates in a periodic manner. Hes7 itself then functions as a transcriptional repressor, leading to negative feedback that contributes to its oscillatory expression pattern.
- FGF (Fibroblast Growth Factor) Signaling: FGF signaling, which is high in the posterior PSM, is also involved in the regulation of oscillatory genes. FGF maintains cells in an undifferentiated state in the posterior PSM and influences the periodicity of oscillatory genes. It works in conjunction with Wnt signaling to sustain the oscillatory behavior in the posterior PSM, and as cells progress anteriorly (where FGF levels decrease), the oscillations slow and eventually stop, signaling the onset of somite boundary formation.
- Wnt Signaling Pathway: Similar to FGF, Wnt signaling is active in the posterior PSM and forms a gradient that decreases toward the anterior. The Wnt pathway contributes to the maintenance and synchronization of oscillatory gene expression by interacting with Notch and FGF signaling pathways. This pathway also influences genes like Axin2, which oscillates in the PSM and is involved in the regulation of the segmentation clock.
- Negative Feedback Loops: Oscillatory genes, such as Hes7, are regulated by negative feedback mechanisms that are crucial for their periodic expression. Hes7 protein represses its own transcription by binding to its promoter, leading to a delay in re-expression after degradation. This negative feedback loop creates the oscillatory dynamics, with cycles of expression and repression in a tightly regulated timeframe.
- Intercellular Coupling and Synchronization: The synchronization of oscillations between neighboring cells in the PSM is mediated by Notch signaling. Notch allows cells to communicate and coordinate their oscillatory patterns with one another, ensuring a cohesive wave of gene expression that moves across the PSM. This coupling is essential for creating a uniform segmentation pattern along the anterior-posterior axis.
- Clock Genes (such as Lfng): Other clock-related genes, like Lfng (Lunatic fringe), also oscillate in synchrony with Hes7. Lfng modulates Notch receptor sensitivity, adding an additional layer of feedback regulation that contributes to the segmentation clock.
In summary, the oscillatory expression of genes like Hes7 (hairy) is regulated by a complex interplay of Notch, FGF, and Wnt signaling pathways, supported by negative feedback loops and intercellular coupling. These regulatory interactions form the segmentation clock that controls the rhythmic, timed formation of somites in somitogenesis.
(ChatGPT 4o)
Fgf4 maintains Hes7 levels critical for normal somite segmentation clock function Nov 19, 2020 https://doi.org/10.7554/eLife.55608 https://elifesciences.org/articles/55608
15 April 2000 Zebrafish Mesp family genes, mesp-a and mesp-b are segmentally expressed in the presomitic mesoderm, and Mesp-b confers the anterior identity to the developing somites Atsushi Sawada, Andreas Fritz, Yun-Jin Jiang, Akihito Yamamoto, Kyo Yama https://journals.biologists.com/dev/article/127/8/1691/41196/Zebrafish-Mesp-family-genes-mesp-a-and-mesp-b-are
RARβ2 positively regulates Tbx3 a marker of hypaxial muscle, and negatively regulates Tbx6 via Ripply2 to restrict the anterior boundaries of the presomitic mesoderm and caudal progenitor pool. (要旨より)(カエルの実験)
Rostral shifting and expansion of somitomere and presomitic mesoderm markers occurs in Rarβ2 MO-injected embryos. (A-F) Embryos were injected unilaterally at the 2- or 4-cell stage with 26 ng Rarβ2.L MO+26 ng Rarβ2.S MO. Injected side is indicated by magenta β-gal lineage tracer. Neurula stage embryos shown in dorsal view with anterior on the left. Rarβ2 MOs rostrally shift somitomere markers Ripply2 (A) and Mespa/Thyl2 (B), and thicken their boundaries of expression (Ripply2, 25/27 embryos; Mespa, 26/31). The expression domains of presomitic mesoderm markers Tbx6 (C), Msgn1 (D) and Fgf8 (E), and the Notch direct target Esr5 (F) are expanded rostrally (red vertical lines) by Rarβ2 MOs (Tbx6, 26/28 embryos; Msgn1, 7/9; Fgf8, 9/13; Esr5, 19/20). Broken red line indicates the midline.
RARβ2 is required for vertebrate somitogenesis Amanda Janesick, Weiyi Tang, Tuyen T. L. Nguyen, Bruce Blumberg Development (2017) 144 (11): 1997–2008.
- the mechanisms regulating the spatially restricted and periodic expression of Mesp2 have remained elusive
Mesp2 and Tbx6 cooperatively create periodic patterns coupled with the clock machinery during mouse somitogenesis Masayuki Oginuma, Yasutaka Niwa, Deborah L. Chapman, Yumiko Saga Development 01 August 2008
Eph gene expression is indeed driven by Mesp2 during somitogenesis, specifically in the context of establishing somite boundaries and the rostrocaudal (head-to-tail) polarity within each forming somite. Here’s how this relationship works:
- Activation of EphA4 by Mesp2: Mesp2, which is expressed in the anterior portion of each forming somite, directly induces the expression of EphA4, a member of the Eph receptor family. This expression is crucial for defining the boundary between the anterior and posterior portions of the somite and establishing proper segmentation.
- Role in Boundary Formation and Polarity: EphA4, once expressed in the anterior half of a somite under the influence of Mesp2, interacts with ephrin ligands (e.g., ephrin-B2) expressed in the posterior half of the neighboring somite. This Eph-ephrin interaction creates a repulsive boundary that physically separates adjacent somites, ensuring they form as distinct segments.
- Regulation of Rostrocaudal Polarity: By driving EphA4 expression in the anterior portion of each somite, Mesp2 helps establish distinct anterior and posterior identities within each somite. This polarity is essential for correct vertebral patterning and alignment of tissues derived from somites.
In summary, Mesp2 directly activates EphA4 expression, which plays a central role in somite boundary formation and establishing rostrocaudal polarity through interactions with ephrin ligands in adjacent segments. This regulatory pathway ensures that somites form with distinct, polarized boundaries essential for the segmented organization of the vertebrate body plan.
（ChatGPT)
Eph受容体はMesp2により遺伝子発現が誘導されるという上の説明でしたがephrinリガンドに関しては、そのような制御機構はないようです。また別の仕組みがephrinリガンドの遺伝子発現を制御しているとのこと（下の説明）。
Mesp2 does not directly drive ephrin gene expression. Instead, Mesp2 primarily drives the expression of EphA4 in the anterior portion of each forming somite, while ephrin (specifically ephrin-B2) expression is regulated in the posterior region of the somite through other mechanisms.
Here’s how the relationship between Mesp2, Eph receptors, and ephrins functions in somite boundary formation:
- Mesp2-Driven EphA4 Expression: Mesp2 induces the expression of EphA4 specifically in the anterior half of the forming somite. This localized EphA4 expression is crucial for the anterior-posterior patterning and segmentation process.
- Ephrin-B2 Expression in the Posterior: Ephrin-B2 is expressed in the posterior half of the adjacent somite, not directly due to Mesp2 but rather through regulatory mechanisms that establish complementary expression patterns of Eph receptors and ephrins across somite boundaries. The ephrin-B2 gene is expressed in response to other signaling inputs that are distinct from those regulating Mesp2.
- Eph-Ephrin Interaction and Boundary Formation: The interaction between EphA4 (induced by Mesp2 in the anterior half of a somite) and ephrin-B2 (in the posterior half of the adjacent somite) creates a repulsive boundary between somites. This boundary formation does not require Mesp2 to directly regulate ephrin expression; rather, it relies on the complementary expression and interaction of Eph receptors and ephrins across the somite boundary.
In summary, Mesp2 specifically drives the expression of EphA4 but not ephrin genes. The complementary expression of ephrin-B2 in the posterior region of adjacent somites is regulated by other factors, allowing for the necessary Eph-ephrin interaction that leads to somite boundary formation and segmentation.
（ChatGPT)
一つの体節とその直後の体節との境界がどうやってきまるのかというと、「反発シグナル」であるEph（リガンド）/ephrin（受容体）シグナリングが活躍しているそうです。Ephリガンドは体節の尾側の境界になる細胞で発現します。ephrin受容体はその体節の尾側の体節の吻側境界となる細胞に発現します。
Patterning and mechanics of somite boundaries in zebrafish embryos Save Related Papers Chat with paper May 2020Seminars in Cell and Developmental Biology 107(8–10) DOI:10.1016/j.semcdb.2020.04.014 LicenseCC BY-NC-ND 4.0 https://www.researchgate.net/figure/Repulsive-cues-A-Eph-ephrin-molecules-are-expressed-in-stripes-in-the-anterior-PSM_fig2_341505871
The “clock-and-wavefront” hypothesis provides a model to explain the periodic formation and size of somites along the body axis, including how changes in this mechanism can lead to variations in neck length across species, such as the long neck of a giraffe. This model combines two main elements: a segmentation clock (oscillatory genes like Hes) and a wavefront (signaling gradients) that together regulate the number and size of somites, which ultimately determine vertebral length and number. Here’s how this applies to somite length and neck length in species with elongated necks like the giraffe:
- Segmentation Clock and Somite Number:
- In the clock-and-wavefront model, the segmentation clock operates via oscillations of genes like Hes7 in the presomitic mesoderm (PSM), which determine the periodicity of somite formation. The frequency of these oscillations sets the timing for each somite’s formation: a faster clock results in smaller somites, while a slower clock results in larger somites.
- In giraffes, it is not just the somite size but also the number of cervical somites that contributes to their long necks. Giraffes have a greater number of cervical vertebrae due to changes in clock periodicity or regulatory mechanisms that extend the duration of somite formation specifically in the neck region.
- Wavefront Position and Somite Size:
- The wavefront in this model is governed by opposing gradients of FGF/Wnt (high in the posterior PSM) and retinoic acid (RA) (high in the anterior PSM). These gradients set a threshold (the wavefront) where cells transition from an oscillatory state to a stable, segmented state, allowing them to form a somite.
- The speed at which the wavefront moves anteriorly relative to the segmentation clock determines the length of each somite. In species with longer necks, slower wavefront progression or a slower segmentation clock could lead to more numerous and/or larger somites, thus lengthening the neck region.
- Genetic and Evolutionary Adaptations in Giraffes:
- In giraffes, genetic modifications likely adjust the clock and wavefront to generate more and/or slightly larger cervical somites compared to shorter-necked animals. Evolutionary adaptations may involve changes in the expression or function of segmentation genes (Hes7, Lfng) or wavefront-controlling signals (FGF/Wnt and RA pathways) specifically in the neck region, resulting in an extended period of somite formation.
- This extended segmentation in the cervical region leads to a greater number of vertebrae in the neck, thereby lengthening it. This mechanism can work similarly in other animals with elongated body segments, such as snakes, where a higher number of smaller somites along the entire body axis results in a long, flexible body.
- Somite Length and Scaling:
- The size of each somite is largely determined by the interplay between the oscillation period of the segmentation clock and the speed of the wavefront. Slower oscillations and/or a more gradual wavefront progression allow each somite to form at a larger size. Conversely, faster oscillations and a rapid wavefront progression create smaller somites.
- In giraffes, modifications in these parameters during embryonic development result in relatively longer somites in the cervical region, though the exact cellular and molecular adjustments in giraffes are still an area of research.
In summary, the clock-and-wavefront hypothesis explains the length of the giraffe’s neck through modifications in the timing of the segmentation clock, the movement of the wavefront, and the regional regulation of somite number and size. Together, these factors lead to a greater number and/or larger size of cervical somites, contributing to the giraffe’s unique neck length.
(ChatGPT 4o)
The “determination front” in somitogenesis is generated by the interaction and balance between opposing gradients of retinoic acid (RA) and FGF/Wnt signaling within the presomitic mesoderm (PSM). This front serves as a threshold boundary where cells transition from an oscillatory state to a stable, segmented state, leading to the formation of somites. Here’s how the determination front is generated and how cells interpret the gradient ratios:
1. Generation of the Determination Front
The determination front is established by:
- Opposing Gradients:
- FGF and Wnt signaling are highly active in the posterior PSM, where they promote cell proliferation and maintain cells in an undifferentiated, mesenchymal, and oscillatory state.
- Retinoic acid (RA), produced in the anterior PSM, creates a gradient that opposes FGF and Wnt, promoting differentiation and segmentation.
- Anterior-Posterior Positioning:
- Cells in the PSM are sequentially positioned along the anterior-posterior axis as the embryo elongates, so as cells move closer to the anterior, they encounter gradually decreasing levels of FGF and Wnt and increasing levels of RA.
- The determination front forms where these gradients reach a critical balance. This is where the FGF/Wnt signals are low enough, and RA is high enough, to trigger the transition from oscillatory to stabilized gene expression, allowing segmentation to occur.
2. Interpretation of the RA and FGF/Wnt Gradient Ratios
Cells in the PSM interpret the ratio of RA to FGF/Wnt, which serves as a positional cue:
- Threshold Mechanism:
- Cells use the relative levels of FGF/Wnt and RA to determine whether they are within the oscillatory region (posterior) or the segmentation-competent region (anterior).
- When FGF/Wnt signals are high relative to RA, cells maintain the oscillatory gene expression pattern (segmentation clock).
- When RA levels become higher than FGF/Wnt at the determination front, the oscillations stop, and cells enter a segmentation-competent state. This ratio effectively serves as a “threshold” that cells interpret to decide when to commit to somite formation.
- Effect on Oscillatory Gene Expression:
- High FGF and Wnt signaling maintains the expression of oscillatory genes like Hes7, which are essential for the segmentation clock’s periodic cycles.
- As cells move anteriorly and the RA/FGF-Wnt ratio increases, oscillatory gene expression is gradually suppressed. This stabilization in gene expression coincides with the expression of segmentation-specific genes like Mesp2 at the determination front, which marks the start of somite boundary formation.
- Gradual Transition from Oscillatory to Stable State:
- The transition from an oscillatory to stable gene expression state happens progressively as cells move across the determination front.
- This ratio of RA to FGF/Wnt ensures that segmentation occurs at the correct position, as cells “sense” this gradient ratio and stop oscillating only when they reach the front.
Summary
The determination front is generated by the opposing RA and FGF/Wnt gradients, which establish a positional threshold in the PSM. Cells interpret the ratio of RA to FGF/Wnt as a signal to either maintain oscillatory gene expression (when FGF/Wnt is high) or transition to a stable state for somite formation (when RA is high). This ratio serves as a positional cue, enabling cells to transition to segmentation precisely at the right position along the axis, ensuring orderly somite formation.
（ChatGPT 4o)
80451-1/asset/f64e0663-0e25-4a61-9609-9f095219f249/main.assets/gr3_lrg.jpg)
