約 2,885,860 件
https://w.atwiki.jp/morrowind/pages/437.html
【俺Mod】Dungeon Phobia【屋内でテレポートすると死ぬ】 最終更新日 2009-12-13 タグ #D *AzuMite ゲームシステム マゾ向け 俺Mod Dungeon Phobia ダンジョン恐怖症 Version 1.01 (2009-12-13) Author AzuMite http //www24.atwiki.jp/morrowind/ 概要 Interior Cellでテレポート系の魔法か薬かスクロールか魔法の道具を使うと大きなペナルティーを受ける。目的はダンジョンの奥深くで死にそうになっても安易にテレポートで脱出できないようにすることで、ダンジョン探索の緊張感を高めること。 ペナルティーは死。これはやりすぎかもしれないので将来のバージョンで変更する可能性がある。 インストール 普通に。 衝突するMod 次のスクリプトを変更するMod。 amuletAundaeScript amuletBerneScript amuletQuarraScript ゲームの途中から導入しても大丈夫か 大丈夫だと思う。 詳細 下記の効果を持つ魔法か薬かスクロールか魔法の道具をInterior Cellで使用すると、テレポート直後に即死する。 Almsivi Intervention Divine Intervention Recall 吸血鬼が自分の血族の本拠地にテレポートできる護符をInterior Cellでで使うと、テレポート直後に即死する。 屋外に見せかけたInterior Cellからテレポートすると死ぬ。しかし特殊なActivator「am_dp_ac_allowteleport」が設置されたセルからは安全にテレポートできる。Mournholdの街は対応済み。他のModで追加される屋外に見せかけたInterior Cellには対応できない。 Cell "Ministry of Truth, Holding Cells"はInterior Cellだが安全にテレポート可能。メインクエスト絡み。 屋内にいるのか屋外にいるのかのチェックは30フレーム毎に行なうので、屋外へ出た直後にテレポートしようとすると死ぬかも。 その他 仕組みは可能な限り単純にした。複雑にする程バグが入り込む余地ができるので。 本来ならDisableTeleporting関数を使うべきなのだが、関数が壊れているのかどうやってもエラーを吐くので諦めた。 PCがテレポートするのを阻止する方法を思いつかなかったので、テレポートしたら死ぬことにした。 更新履歴 1.0 (2009-11-29) 最初期版 1.01 (2009-12-13) 吸血鬼が自分の血族の本拠地にテレポートできる護符のスクリプトに手を入れて、屋内で使うと死ぬようにした。 ダウンロード このページにアップロードされたファイル -- Dungeon Phobia 101.zip コメント欄 1.01 -- 管理人 (2009-12-13 13 16 51) 名前 コメント
https://w.atwiki.jp/nicepaper/pages/101.html
CSSで文字やテキストを上下中央の中心揃えにする方法について書いていきます。6月20日記事 イメージ display table-cellで中央に ブロック要素を中央に 目次 このサンプルコードをいじっていきます。 display table-cellで上下中央揃えしてみる。 ブロック要素の方も試してみる。 このサンプルコードをいじっていきます。 HTML部 !DOCTYPE HTML html head meta charset="utf-8" link rel="stylesheet" type="text/css" href="reset.css" link rel="stylesheet" type="text/css" href="style.css" title 上下中央揃え /title /head body div id="wrapper" div id="box" p 文字が入っています。これを中央にするためには br どのようにすればよいのでしょうか？ br 考えていきます。 /p /div /div /body /html CSS部 @charset "utf-8"; /* CSS Document */ #wrapper{ margin 10px auto; padding 20px; width 800px; height 400px; border 1px solid #000; } #box{ margin 30px auto; width 600px; height 300px; border 1px solid #000; } p{ } すると次のような表示になります。 paddingを設定しているため、#boxのmargin-topが効いています。 ※CSSでmarginプロパティが効かない時の対策を参照 display table-cellで上下中央揃えしてみる。 display table-cell;vertical-align middle;text-align center;を追加していきます。 CSS部 @charset "utf-8"; /* CSS Document */ #wrapper{ margin 10px auto; padding 20px; width 800px; height 400px; border 1px solid #000; } #box{ margin 30px auto; width 600px; height 300px; border 1px solid #000; display table-cell; vertical-align middle; text-align center; } p{ } すると次のような表示になります。 vertical-alignは インライン要素かtable-cellしかききません！ marginが効かなくなり、左上にボックスが配置されますが、文字が上下中央に揃います。 適宜ボックス等をmargin,padding,borderを使って動かないよう固定化し、上下中央を行ってください。 ブロック要素の方も試してみる。 #boxにposition relativeを追加し、p要素の方にブロックを中央にさせるコードを記載していきます。多少めんどくさいですが、次のように左右上下からの位置を0としmargin autoをすると上下中央になります。 @charset "utf-8"; /* CSS Document */ #wrapper{ margin 10px auto; padding 20px; width 800px; height 400px; border 1px solid #000; } #box{ margin 30px auto; width 600px; height 300px; border 1px solid #000; position relative; } p{ position absolute; top 0; left 0; right 0; bottom 0; margin auto; width 200px; height 100px; background #888; } このようになりました。 ブロック要素のほうは多少コードが増えてめんどくさいですが、これでも上下中央にすることができます。 上下中央にさせるときにブロックか文章かを考えておくと切り分けが楽になると思います。 以上
https://w.atwiki.jp/jasagiri/pages/54.html
http //blog.lawrencepit.com/2008/06/18/hello-functor-pattern-based-method-dispatch-in-ruby/ Hello Functor pattern based method dispatch in ruby June 18, 2008 ruby 界隈で一歩先へいく革新的なイニシアティブのひとつがWavesプロジェクトです。 これはrubyベースのウェブアプリケーションでオープンソースフレームワークです。 これは皆が「次世代」だと思うはずです。 これは美しく簡潔なコードで書かれています。 Lambdas はwavesでは非常にはっきりしています。 リソースの実装が マッピングを基礎付けました。 MVC パラダイム (せいぜい必要な)を調べ始める前にResources は一級市民です（？） このプロジェクトからデザインと実装の美しい2つの小さなgemが生まれました。 1つは Autocodeで 、クラスとモジュールを自動的に読み込みんだり生成したりするライブラリです。 Autocode に関する詳しい情報がここ（http //dev.zeraweb.com/introducing-autocode-0-9-9/）にあります。 もうひとつは、私にとってより魅力的なもので Functor と呼ばれるシンプルで小さなgemです。それはここで詳しく説明します。これは Topher Cyll 氏が最近開発した multi gem の成果を元に作られています。 Waves プロジェクトからは独立した Functor の使用例。： functor( normalize, Symbol, Hash ) { | name, options | options.merge!( name = name ) } functor( normalize, String, Hash ) { | p, options | options.merge!( pattern = p ) } functor( normalize, Regexp, Hash ) { | re, options | options.merge!( pattern = re ) } functor( normalize, Exception ) { | exception | { exception = exception } } functor( normalize, Hash ) { | options | options } これはこうやって呼ばれます。： def map( *args, block ) options = normalize( *args ) ...end このコードは Functor 無しでどのようになるか考えてみてください。多分1ダースの入れ子の if-then-else や case-when などの構文になるでしょう。 さて、Functor はメソッドを多重定義するように振舞うように見えます。 java や C++ のような言語にはネイティブで存在しています。 Matz は「直交した特徴 [...] は複雑で爆発できる」(？)ため、ruby にメソッドの多重定義を実装するのを拒んでいます。 私は彼が正しいと思います。 Functor は直交性をこっそり許します。でもスタッフと一緒に船から落ちないでください。 あなたは正しいのですから。（？） Functor の美しさはメソッドの多重定義以上のことを許容します。メソッドの多重定義することはクラスの引数の議論です。対照的に Functor は引数の引き当てにまず === 演算子を試し、それから == 演算子、そしてそれでもマッチしなかったら、あなたが提供している lambda や Proc（または何もあたらなければ call が返されます） を試します。 このためこんなことが可能になるでしょう。： functor( get, /^foo$/ ) { |p| "foo!" } functor( get, /^bar$/ ) { |p| "bar!" } get("bar") を呼ぶと bar! が返ります あなたはfunctorをgemでインストールして弄れます (source is http //gems.github.com/) sudo gem install -r functor または ソースコードは github で手に入れることができます。
https://w.atwiki.jp/killingbeller/pages/26.html
Bell s Cup 1.zip info 【大会名】 Bell s Cup 【大会の主旨】 大会開催のテスト運営 兼 特殊制限下での遊戯王環境のシミュレート目的。 【対戦形式】 マッチ(人数が多い場合予選はシングル) 【開催日時】 2013年4月2日(火) 11 50参加締切 12 00～開催予定。 【参加方法】 大会用のhamachiのネットワークに参加 【部屋名とパスワード】 ●ネットワーク名● KB-0402-01 KB-0402-02 KB-0402-03 KB-0402-04 ●ネットワークパスワード● 0402（上記の全ネットワーク共通） 【備考・その他】 適用制限のlflistは同フォルダ内にあります。 【質問等の受付先】 http //killingbeller.blog.fc2.com/blog-entry-19.html 上記URLもしくは大会用のhamachiネットワークからのチャットでも可。 【禁止制限】 ●特殊制限ルール● エクストラデッキに同名カードは複数積みできない。 魔法カード・罠カード・手札誘発の効果モンスターカードはデッキに３積みできない。 ●追加禁止カード● ５枚 《甲虫装機 ホーネット》69207766 《Ｎｏ.１１ ビッグ・アイ》 80117527 《グリモの魔導書》89739383 《魔導書の神判》46448938 《血の代償》80604091 ●追加制限カード● ２０枚 《聖なる魔術師》31560081 《デビル・フランケン》69015963 《サウザンド・アイズ・サクリファイス》63519819 《発条空母ゼンマイティ》 81122844 《蝶の短剣－エルマ》69243953 《トラゴエディア》98777036 《大天使クリスティア》59509952 《光と闇の竜》47297616 《水精鱗－メガロアビス》21954587 《マシンナーズ・フォートレス》05556499 《神光の宣告者》44665365 《王立魔法図書館》70791313 《暗黒界の術師 スノウ》60228941 《炎星師－チョウテン》01662004 《創造の代行者 ヴィーナス》64734921 《リチュアの儀水鏡》46159582 《封印の黄金櫃》75500286 《簡易融合》01845204 《激流葬》53582587 《聖なるバリア－ミラーフォース》44095762 ●追加準制限カード● ５枚 《裁きの龍》57774843 《カオス・ソーサラー》09596126 《馬頭鬼》92826944 《ローンファイア・ブロッサム》48686504 《深海のディーヴァ》78868119 《光の援軍》94886282 ●追加無制限カード● ５枚 《甲虫装機 ダンセル》68184115 《神秘の代行者 アース》91188343 《グローアップ・バルブ》67441435 《月読命》34853266 report Bell s Cup （2013/04/02 12 00頃 開催） 大会の詳細情報は『BC-0402_info.txt』または以下のURLに載せています。 （http //killingbeller.blog.fc2.com/blog-entry-20.html） 今回の参加者は以下の６名となります（敬称略） whiter ：【兎カラクリ】 ygo!magmag ：【甲虫装機】 練習 ：【ヴァイロン】 むらさきサイクロン：【HEROビート】 wataV ：【サンダー】 kui ：【兎ヴェルズ】 抽選の結果、以下のプログラムになりました。 人数の関係上、kuiさんとwhiterさんはシード枠になりました。 第一回戦：むらさきサイクロン vs wataV 先攻○ ×後攻 後攻× ○先攻 先攻○ ×後攻 第二回戦：ygo!magmag vs 練習 先攻○ ×後攻 後攻× ○先攻 先攻○ ×後攻 第三回戦(準決勝)：kui (シード枠) vs むらさきサイクロン 先攻○ ×後攻 後攻○ ×先攻 第四回戦(準決勝)：ygo!magmag vs whiter (シード枠) 先攻○ ×後攻 後攻× ○先攻 先攻○ ×後攻 第五回戦(決勝)：kui vs ygo!magmag 先攻× ○後攻 先攻○ ×後攻 後攻× ○先攻 第六回戦(３位決定戦)：むらさきサイクロン vs whiter 先攻○ ×後攻 後攻○ ×先攻 よって結果は以下の順位となりました。 １位：ygo!magmag ２位：kui ３位：むらさきサイクロン 参加者の皆様、お疲れ様でした。 ご参加ありがとうございました。 Beller
https://w.atwiki.jp/emcs/pages/22.html
書式 (setcar CELL NEWCAR) 第1引数のコンスセルのcarをNEWCARに置き換える。 戻り値はNEWCAR。 (setq cell '(a b)) #= (a b) cell #= (a b) (setcar cell 'z) #= z cell #= (z b) 名前 コメント
https://w.atwiki.jp/hogazurou/pages/25.html
リンク 課題ページ lab3A(１つ目) #include stdio.h #include stdlib.h #define LISTNUM 6 void stack(); void que(); struct cell{//cellの構造体 int element; struct cell *next;//次のcellを指すポインタ }; int main(int argc, char* argv[]) { int celect; printf("stack 1\nque 0\nboth else\n"); scanf("%d", celect); if(celect==1) stack(); else if(celect==0) que(); else if(celect!=1 celect!=0) { stack(); que(); } return 0; } void stack(){ printf("input\n"); int a,b; struct cell *p,*first; first=(cell *)malloc(sizeof(cell)); first- next=NULL; for(a=0;a LISTNUM;a++){ scanf("%d", b); p=(cell *)malloc(sizeof(cell)); p- element=b; p- next=first- next; first- next=p; } printf("stack "); p=first- next; while(p!=NULL){ printf("%d→",p- element); p=p- next;//次のセルへ移動 } printf("\n"); } void que(){ printf("input\n"); int a,b; struct cell *p,*q,*first; first=(cell *)malloc(sizeof(cell)); first- next=NULL; q=NULL; for(a=0;a LISTNUM;a++){ scanf("%d", b); p=(cell *)malloc(sizeof(cell)); p- element=b; p- next=NULL; if(first- next==NULL){ first- next=p; } else { q- next=p; } q=p; } printf("que "); p=first- next; while(p!=NULL){ printf("%d→",p- element); p=p- next; } printf("\n"); } lab3B(プログラム) #include stdio.h void push(int a,int i); void pop(int); void deque(int); int a[6];//関数内でも同じ数字を操作するためにメインの外に書いた int main(int argc,char *argv[]){ int c,i; for(i=0;i 6;i++){ scanf("%d", c);//取り込んだ数字を push(c,i);//順番にpushする } printf("stack "); while(i 0){//最後に入れたものから順に i--;//数字を減らしていき pop(i);//出力する } printf("\nque"); while(i 6){//最初に入れたものから順に deque(i);//出力する i++; } return 0; } //配列i番目にbを代入する関数 void push(int b,int i){ a[i]=b; } //配列i番目をprintする関数 void pop(int i){ printf("%d",a[i]); } //上と同じ void deque(int i){ printf("%d",a[i]); } いまいちなできですが、表示的にはＯＫのはず。
https://w.atwiki.jp/mtgflavortext/pages/1000.html
暗く湿った地下室を避ける他の理由が必要かのような。 As if anyone needed another reason to avoid dark, dank cellars. イニストラード 【M TG Wiki】 名前
https://w.atwiki.jp/geckjp/pages/15.html
このページでは、初めてGECKを使う人向けに自分の家をつくる方法を説明します。 1.マスターファイルを読み込もう GECKを立ち上げて、まず始めにやるのはマスターファイルを読み込む事です。フォルダのアイコンをクリックしてDataウィンドウを出し、fallout.esmを読み込んでください。 マスターファイルとは、Fallout3の内部データ（NPC、アイテム、MAP、クエストなど全てのゲームデータ）が入っているファイルです。 GECKでは、このファイルに変更を加えていって、差分データをプラグイン(拡張子esp)として保存するため、マスターファイルが壊れてゲームができなくなるということはないので、心配無用です。 2回目からの編集では保存したプラグインファイルを選択し、Set as Active Fileボタンを押して編集状態にしなければいけません。 それと、2回目からはマスターファイルは選択する必要が無いです。（プラグインにマスターファイルも同時に読み込むと設定されているため） 2.家のCellを作ろう Cellとは、簡単に言うととても大きい部屋のようなものです。モイラの雑貨店、テンペニータワーのロビーなど、屋内（Interior）の場所は独立したCellにその場所のデータが入っています。 自分の家の内部のCellを作るために上のメニューからWorld Cellと進んで、World Spaceの欄をInteriorにし、他のCellの名前がずらずら並んでいるところで右クリックをし、Newを押します。 すると、新しいIDを入力しろと言われるので、「aaorenoie」などと分かりやすい名前でつけておいてください。 名前の最初にaaとつける理由は、ID順で一番最初に出てきてやりやすいのでです。 3.家の内部を作ろう 家の内部を編集するために、今作ったCellをRender Windowに表示させます。 CellViewから今作ったCellをダブルクリックしてください。何も無い空間がRendarWindowに表示されます。 この何も無い空間に物を配置していきます。 ObjectWindowからStatic（静的オブジェクト）のカテゴリを選ぶとオブジェクトのリストが表示されます。 とりあえずベッドや椅子よりも先にガワを配置した方がいいので、Filterに「WoodShackInterior」と入力し、右側のリストに表示されたWoodShackInteriorをRenderWindowにドラッグ ドロップしてください。すると汚い小屋の屋内が現れます。（真っ黒の物がでてきたよShit！って人は上のランプのアイコンをクリックしてください） なんでこの汚い小屋をわざわざ自分の家にしたかというと、これ以外の家の内部のオブジェクトはパズルピースのようになっておりコツが必要だからです。
https://w.atwiki.jp/attery/pages/2.html
Working Principle and Uses ry cell battery was being developed in the year 1887 by a scientist named Karl Gassner of German origin. This device was later patented in the year 1982. Before we begin with understanding the composition and working of a dry cell battery, it is important to understand what a battery is. A battery is a device or an instrument that yields electricity by undergoing certain chemical reactions. Unlike the wet cell batteries, the dry cell batteries are non rechargeable. The dry cell battery as the name suggests doesn t carry any type of liquid. Instead it contains a paste which acts as the electrolyte. The cell battery consists of a paste because it is thicker in consistency and thus, will not spill. This battery electrolyte contains little moisture, just enough to allow current to flow through it. Read further to know more about the dry cell battery in detail. How Does a Dry Cell Battery Work?http //www.toshibaprobe.com/MF1B54L002.html Reusable Spo2 Sensors The role of any battery is to convert chemical energy to electrical energy. Dry cell battery is a voltage producing battery, containing the electrolyte chemical in the form of a thick paste. It produces voltage of about 1.5 volts. According to science, an electrode is a conductor used in a battery to run the circuit. It can have positive or negative charge. In the above diagram (which represents the parts of a dry cell battery), the zinc casing provides enclosing to the electrolyte and the cathode as a whole. Ammonium chloride is the electrolyte used in this type of battery. Just ahead of the electrolyte lies manganese dioxide separated from the electrolyte by a separator or partitioning. At the center of the cell battery lies a carbon rod, which is the cathode. Zinc casing is considered as the anode. Now that we know about the structure and parts of the dry cell battery, let us understand its working. Every portion of the battery device has chemical reactions occurring in it. While a reduction reaction occurs at the carbon electrode, oxidation occurs at the zinc casing, which is the anode. The chemical reactions that define the entire energy generation process are as follows The ammonium ion from ammonium chloride reacts with two electrons to produce ammonia and hydrogen gas.
https://w.atwiki.jp/tiger/pages/4.html
TEC-/- BTK-/- double mutant T cells exhibit severely impaired T cell activitity. RLK-/-ITK-/- double mutant celles exhibit severely imparired Th2 responses. Grb2(+/-) mice disrupt T cell signaling networks and development. Dendric cells and macrophages of MEK3 deficient mice have impaired IL12 production. Bam32(-/-) B cell develop normally but have impaired T-independent antibody responses in vivo. T-cell and B-cell of RAP1A deficient mice impair integrin-mediated cell adhesion. T-cell of WASP deficient mice impair the proliferaction and antigen receptor cap formation in response to anti-CD3zeta stimulation. T-cell of SHB defective mice impair the phosphorylation of LAT and consequently the activation of MAP kinase pathways. B-cell of 3BP2 (-/-) deficient mice have defective in proliferation, cell cycle progression, PLC-gamma2 phosphorylation, calcium mobilization, NF-ATp dephosphorylation, and Erk and Jnk activation in response to BCR ligation. B-cell of Vav2(-/-) deficient mice are defective in the ability to switch immunoglobulin class. T-cell of Vav1(-/-) deficient mice exhibit impaired antigen receptor signaling. Vav1(-/-)Vav2(-/-) mice exhibit greatly reduced the mature B-cells. Vav-1-/-Vav-2-/- B cells were unresponsive to BCR-driven proliferation in vitro and to thymus-indepen-dent antigen in vivo. Fyn-deficient mice exhibit a remarkably specific lymphoid defect thymocytes are refractile to stimulation through the TCR with mitogen or antigen. Lck-deficient mice show a pronounced thymic atrophy, with a dramatic reduction in the double-positive (CD4+CD8+) thymocyte population. T cell from mice deficient in LCK is required for normal signal transduction through the TCR. T cells from mice deficient in SLAP-130/Fyb show markedly impaired proliferation. B cell of chicken deficient ITK reduce IP3 generation and phospholipase C gamma 2 tyrosine phosphorylation. T cell of mice deficient ITK reduce IP3 generation and phospholipase C gamma 1 tyrosine phosphorylation. T cell of mice deficient ITK have failure of Th2 development. Mice deficient in ITK have reduced proliferative responses to MHC stimulation and to anti-TCR cross-linking Mutations in Btk cause X-linked immunodeficiency. Gads(GRAP2) has a role in thymocyte proliferaction for maturation of T-cells. Gads(GRAP2) has a role for homeostatic proliferaction in B cells. Grap negatively regulates T-cell proliferation. Gab2 is a substrate of ZAP-70 and functions as a switch molecule toward inhibition of TCR signal transuduction. B cell signaling causes tyrosine phosphorylation of Gab1, and in turn SHP2 bind to Gab1 Gab1 phosophorylation potentiate the phosphorylation of Akt, PI3K-dependent response. RasGRP1 mediates Ras activation following TCR stimulatioin. RasGRP1 and RasGRP3 induces RAS activation in B-cell to response to T-cell stimulation. Grb2-hSos1-PLCgamma1-p36/p38-ZAP70 complexes localize in the vicinity of TCR-zeta Gads(Grap2) plays an important role in T-cell signaling via its association with SLP-76 and LAT. Lck is required for normal signal transduction through the TCR. ZAP-70 plays crucial roles in T-cell activation and development. Syk triggers cellular activation in T-cell. TEC-/- BTK-/- double mutant T cells exhibit severely impaired T cell activitity. 1 J Exp Med. 2000 Dec 4;192(11) 1611-24. Severe B cell deficiency in mice lacking the tec kinase family members Tec and Btk. Ellmeier W, Jung S, Sunshine MJ, Hatam F, Xu Y, Baltimore D, Mano H, Littman DR. Molecular Pathogenesis Program, Skirball Institute of Biomolecular Medicine. wilfried.ellmeier@univie.ac.at The cytoplasmic protein tyrosine kinase Tec has been proposed to have important functions in hematopoiesis and lymphocyte signal transduction. Here we show that Tec-deficient mice developed normally and had no major phenotypic alterations of the immune system. To reveal potential compensatory roles of other Tec kinases such as Bruton's tyrosine kinase (Btk), Tec/Btk double-deficient mice were generated. These mice exhibited a block at the B220(+)CD43(+) stage of B cell development and displayed a severe reduction of peripheral B cell numbers, particularly immunoglobulin (Ig)M(lo)IgD(hi) B cells. Although Tec/Btk(null) mice were able to form germinal centers, the response to T cell-dependent antigens was impaired. Thus, Tec and Btk together have an important role both during B cell development and in the generation and/or function of the peripheral B cell pool. The ability of Tec to compensate for Btk may also explain phenotypic differences in X-linked immunodeficiency (xid) mice compared with human X-linked agammaglobulinemia (XLA) patients. Publication Types Research Support, Non-U.S. Gov't PMID 11104803 [PubMed - indexed for MEDLINE] RLK-/-ITK-/- double mutant celles exhibit severely imparired Th2 responses. 1 Nat Immunol. 2001 Dec;2(12) 1183-8. Mutation of Tec family kinases alters T helper cell differentiation. Schaeffer EM, Yap GS, Lewis CM, Czar MJ, McVicar DW, Cheever AW, Sher A, Schwartzberg PL. National Human Genome Research Institute, National Institutes of Health, Bethesda, MD 20892, USA. The Tec kinases Rlk and Itk are critical for full T cell receptor (TCR)-induced activation of phospholipase C-gamma and mitogen-activated protein kinase. We show here that the mutation of Rlk and Itk impaired activation of the transcription factors NFAT and AP-1 and production of both T helper type 1 (TH1) and TH2 cytokines. Consistent with these biochemical defects, Itk-/- mice did not generate effective TH2 responses when challenged with Schistosoma mansoni eggs. Paradoxically, the more severely impaired Rlk-/-Itk-/- mice were able to mount a TH2 response and produced TH2 cytokines in response to this challenge. In addition, Rlk-/-Itk-/- cells showed impaired TCR-induced repression of the TH2-inducing transcription factor GATA-3, suggesting a potential mechanism for TH2 development in these hyporesponsive cells. Thus, mutations that affect Tec kinases lead to complex alterations in CD4+ TH cell differentiation. Publication Types Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. PMID 11702066 [PubMed - indexed for MEDLINE] Grb2(+/-) mice disrupt T cell signaling networks and development. 1 Nat Immunol. 2001 Jan;2(1) 29-36. Disruption of T cell signaling networks and development by Grb2 haploid insufficiency. Gong Q, Cheng AM, Akk AM, Alberola-Ila J, Gong G, Pawson T, Chan AC. Howard Hughes Medical Institute, Washington University School of Medicine, St. Louis, MO 63110, USA. The developmental processes of positive and negative selection in the thymus shape the T cell antigen receptor (TCR) repertoire and require the integration of multiple signaling networks. These networks involve the efficient assembly of macromolecular complexes and are mediated by multimodular adaptor proteins that permit the functional integration of distinct signaling molecules. We show here that decreased expression of the adaptor protein Grb2 in Grb2+/- mice weakens TCR-induced c-Jun N-terminal kinase (JNK) and p38, but not extracellular signal-regulated kinase (ERK), activation. In turn, this selective effect decreases the ability of thymocytes to undergo negative, but not positive, selection. We also show that there are differences in the signaling thresholds of the three mitogen-activated protein kinase (MAPK) families. These differences may provide a mechanism by which quantitative differences in signal strength can alter the balance of downstream signaling pathways to induce the qualitatively distinct biological outcomes of proliferation, differentiation or apoptosis. PMID 11135575 [PubMed - indexed for MEDLINE] Dendric cells and macrophages of MEK3 deficient mice have impaired IL12 production. 1 EMBO J. 1999 Apr 1;18(7) 1845-57. Defective IL-12 production in mitogen-activated protein (MAP) kinase kinase 3 (Mkk3)-deficient mice. Lu HT, Yang DD, Wysk M, Gatti E, Mellman I, Davis RJ, Flavell RA. Howard Hughes Medical Institute and Section of Immunobiology, Yale University School of Medicine, New Haven, CT 06520, USA. The p38 mitogen-activated protein kinase (MAPK) pathway, like the c-Jun N-terminal kinase (JNK) MAPK pathway, is activated in response to cellular stress and inflammation and is involved in many fundamental biological processes. To study the role of the p38 MAPK pathway in vivo, we have used homologous recombination in mice to inactivate the Mkk3 gene, one of the two specific MAPK kinases (MAPKKs) that activate p38 MAPK. Mkk3(-/-) mice were viable and fertile; however, they were defective in interleukin-12 (IL-12) production by macrophages and dendritic cells. Interferon-gamma production following immunization with protein antigens and in vitro differentiation of naive T cells is greatly reduced, suggesting an impaired type I cytokine immune response. The effect of the p38 MAPK pathway on IL-12 expression is at least partly transcriptional, since inhibition of this pathway blocks IL-12 p40 promoter activity in macrophage cell lines and IL-12 p40 mRNA is reduced in MKK3-deficient mice. We conclude that the p38 MAP kinase, activated through MKK3, is required for the production of inflammatory cytokines by both antigen-presenting cells and CD4(+) T cells. PMID 10202148 [PubMed - indexed for MEDLINE] Bam32(-/-) B cell develop normally but have impaired T-independent antibody responses in vivo. 1 Immunity. 2003 Oct;19(4) 621-32. Bam32 links the B cell receptor to ERK and JNK and mediates B cell proliferation but not survival. Han A, Saijo K, Mecklenbrauker I, Tarakhovsky A, Nussenzweig MC. Laboratory of Molecular Immunology, The Rockefeller University, New York, NY 10021, USA. Bam32 is an adaptor protein recruited to the plasma membrane upon B cell receptor (BCR) crosslinking in a phosphoinositol 3-kinase (PI3K)-dependent manner; however, its physiologic function is unclear. To determine its physiologic function, we produced Bam32-deficient mice. Bam32(-/-) B cells develop normally but have impaired T-independent antibody responses in vivo and diminished responses to BCR crosslinking in vitro. Biochemical analysis revealed that Bam32 acts in a novel pathway leading from the BCR to MAPK/ERK Kinases (MEK1/2), MAPK/ERK Kinase Kinase-1 (MEKK1), extracellular signal-regulated kinase (ERK), and c-jun NH2-terminal kinase (JNK), but not p38 mitogen-activated protein kinase (p38). This pathway appears to be initiated by hematopoietic progenitor kinase-1 (HPK1), which interacts directly with Bam32, and differs from all previously characterized BCR signaling pathways in that it is required for normal BCR-mediated proliferation but not for B cell survival. PMID 14563325 [PubMed - indexed for MEDLINE] T-cell and B-cell of RAP1A deficient mice impair integrin-mediated cell adhesion. 1 Mol Cell Biol. 2006 Jan;26(2) 643-53. Rap1A-deficient T and B cells show impaired integrin-mediated cell adhesion. Duchniewicz M, Zemojtel T, Kolanczyk M, Grossmann S, Scheele JS, Zwartkruis FJ. Department of Computational Molecular Biology, Max Planck Institute for Molecular Genetics, Ihnestrasse 73, D-14195 Berlin, Germany. Studies in tissue culture cells have demonstrated a role for the Ras-like GTPase Rap1 in the regulation of integrin-mediated cell-matrix and cadherin-mediated cell-cell contacts. To analyze the function of Rap1 in vivo, we have disrupted the Rap1A gene by homologous recombination. Mice homozygous for the deletion allele are viable and fertile. However, primary hematopoietic cells isolated from spleen or thymus have a diminished adhesive capacity on ICAM and fibronectin substrates. In addition, polarization of T cells from Rap1-/- cells after CD3 stimulation was impaired compared to that of wild-type cells. Despite this, these defects did not result in hematopoietic or cell homing abnormalities. Although it is possible that the relatively mild phenotype is a consequence of functional complementation by the Rap1B gene, our genetic studies confirm a role for Rap1A in the regulation of integrins. PMID 16382154 [PubMed - indexed for MEDLINE] T-cell of WASP deficient mice impair the proliferaction and antigen receptor cap formation in response to anti-CD3zeta stimulation. 1 Immunity. 1998 Jul;9(1) 81-91. Wiskott-Aldrich syndrome protein-deficient mice reveal a role for WASP in T but not B cell activation. Snapper SB, Rosen FS, Mizoguchi E, Cohen P, Khan W, Liu CH, Hagemann TL, Kwan SP, Ferrini R, Davidson L, Bhan AK, Alt FW. Howard Hughes Medical Institute, Children's Hospital, Boston, Massachusetts 02115, USA. The Wiskott-Aldrich syndrome (WAS) is a human X-linked immunodeficiency resulting from mutations in a gene (WASP) encoding a cytoplasmic protein implicated in regulating the actin cytoskeleton. To elucidate WASP function, we disrupted the WASP gene in mice by gene-targeted mutation. WASP-deficient mice showed apparently normal lymphocyte development, normal serum immunoglobulin levels, and the capacity to respond to both T-dependent and T-independent type II antigens. However, these mice did have decreased peripheral blood lymphocyte and platelet numbers and developed chronic colitis. Moreover, purified WASP-deficient T cells showed markedly impaired proliferation and antigen receptor cap formation in response to anti-CD3epsilon stimulation. Yet, purified WASP-deficient B cells showed normal responses to anti-Ig stimulation. We discuss the implications of our findings regarding WASP function in receptor signaling and cytoskeletal reorganization in T and B cells and compare the effects of WASP deficiency in mice and humans. PMID 9697838 [PubMed - indexed for MEDLINE] T-cell of SHB defective mice impair the phosphorylation of LAT and consequently the activation of MAP kinase pathways. 9 Sep 24;274(39) 28050-7. Requirement of the Src homology 2 domain protein Shb for T cell receptor-dependent activation of the interleukin-2 gene nuclear factor for activation of T cells element in Jurkat T cells. Lindholm CK, Gylfe E, Zhang W, Samelson LE, Welsh M. Department of Medical Cell Biology, Box 571, Biomedicum, Uppsala University, S-75123 Uppsala, Sweden. Stimulation of the T cell antigen receptor (TCR) induces tyrosine phosphorylation of numerous intracellular proteins. We have recently investigated the role of the adaptor protein Shb in the early events of T cell signaling and observed that Shb associates with Grb2, linker for activation of T cells (LAT) and the TCR zeta-chain in Jurkat cells. We now report that Shb also associates with phospholipase C-gamma1 (PLC-gamma1) in these cells. Overexpression of Src homology 2 domain defective Shb caused diminished phosphorylation of LAT and consequently the activation of mitogen-activated protein kinases was decreased upon TCR stimulation. In addition, the Shb mutant also blocked phosphorylation of PLC-gamma1 and the increase in cytoplasmic Ca(2+) following TCR stimulation. Nuclear factor for activation of T cells is a major target for Ras and calcium signaling pathways in T cells following TCR stimulation, and the overexpression of the mutant Shb prevented TCR-dependent activation of the nuclear factor for activation of T cells. Consequently, endogenous interleukin-2 production was decreased under these conditions. The results indicate a role for Shb as a link between the TCR and downstream signaling events involving LAT and PLC-gamma1 and resulting in the activation of transcription of the interleukin-2 gene. PMID 10488157 [PubMed - indexed for MEDLINE] B-cell of 3BP2 (-/-) deficient mice have defective in proliferation, cell cycle progression, PLC-gamma2 phosphorylation, calcium mobilization, NF-ATp dephosphorylation, and Erk and Jnk activation in response to BCR ligation. 1 Mol Cell Biol. 2006 Jul;26(14) 5214-25. 3BP2 deficiency impairs the response of B cells, but not T cells, to antigen receptor ligation. de la Fuente MA, Kumar L, Lu B, Geha RS. Division of Immunology, Children's Hospital, 300 Longwood Ave., Boston, MA 02115, USA. The adapter protein 3BP2 is expressed in lymphocytes; binds to Syk/ZAP-70, Vav, and phospholipase C-gamma (PLC-gamma); and is thought to be important for interleukin-2 gene transcription in T cells. To define the role of 3BP2 in lymphocyte development and function, we generated 3BP2-deficient mice. T-cell development, proliferation, cytokine secretion, and signaling in response to T-cell receptor (TCR) ligation were all normal in 3BP2(-/-) mice. 3BP2(-/-) mice had increased accumulation of pre-B cells in the bone marrow and a block in the progression of transitional B cells in the spleen from the T1 to the T2 stage, but normal numbers of mature B cells. B-cell proliferation, cell cycle progression, PLC-gamma2 phosphorylation, calcium mobilization, NF-ATp dephosphorylation, and Erk and Jnk activation in response to B-cell receptor (BCR) ligation were all impaired. These results suggest that 3BP2 is important for BCR, but not for TCR signaling. PMID 16809760 [PubMed - indexed for MEDLINE] B-cell of Vav2(-/-) deficient mice are defective in the ability to switch immunoglobulin class. 1 Nat Immunol. 2001 Jun;2(6) 542-7. Comment in Nat Immunol. 2001 Jun;2(6) 482-4. Signal transduction through Vav-2 participates in humoral immune responses and B cell maturation. Doody GM, Bell SE, Vigorito E, Clayton E, McAdam S, Tooze R, Fernandez C, Lee IJ, Turner M. Laboratory of Lymphocyte Signaling and Development, Molecular Immunology Programme, The Babraham Institute, Babraham, Cambridge CB2 4AT, UK. B and T lymphocytes develop normally in mice lacking the guanine nucleotide exchange factor Vav-2. However, the immune responses to type II thymus-independent antigen as well as the primary response to thymus-dependent (TD) antigen are defective. Vav-2-deficient mice are also defective in their ability to switch immunoglobulin class, form germinal centers and generate secondary immune responses to TD antigens. Mice lacking both Vav-1 and Vav-2 contain reduced numbers of B lymphocytes and display a maturational block in the development of mature B cells. B cells from Vav-1(-/-)Vav-2(-/-) mice respond poorly to antigen receptor triggering, both in terms of proliferation and calcium release. These studies show the importance of Vav-2 in humoral immune responses and B cell maturation. PMID 11376342 [PubMed - indexed for MEDLINE] T-cell of Vav1(-/-) deficient mice exhibit impaired antigen receptor signaling. 1 Nature. 1995 Mar 30;374(6521) 474-7. Defective T-cell receptor signalling and positive selection of Vav-deficient CD4+ CD8+ thymocytes. Fischer KD, Zmuldzinas A, Gardner S, Barbacid M, Bernstein A, Guidos C. Program in Molecular Biology and Cancer, Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, Ontario, Canada. During lymphocyte development, cellular proliferation and positive and negative selection events ensure the production of T and B lymphocytes bearing highly diverse, but self-tolerant, repertoires of antigen receptors. These processes are initiated when engagement of growth-factor receptors, or the T and B lymphocyte antigen receptors, induces tyrosine phosphorylation of specific SH2- and SH3-domain-containing cytoplasmic proteins, including Vav. Here we show that vav-/- embryonic stem cells generate only limited numbers of immature and mature T and B lymphocytes in the RAG-2 blastocyst complementation assay. Furthermore, Vav-deficient T lymphocytes showed severely impaired antigen receptor signalling. Finally, we demonstrate that Vav-dependent signalling pathways regulate maturation, but not CD4/CD8 lineage commitment, during T-cell-receptor-mediated positive selection of immature CD4+ CD8+ precursors into mature CD4+ CD8- or CD4- CD8+ T cells. PMID 7700360 [PubMed - indexed for MEDLINE] Vav1(-/-)Vav2(-/-) mice exhibit greatly reduced the mature B-cells. Vav-1-/-Vav-2-/- B cells were unresponsive to BCR-driven proliferation in vitro and to thymus-indepen-dent antigen in vivo. 1 Nat Immunol. 2001 Jun;2(6) 548-55. Comment in Nat Immunol. 2001 Jun;2(6) 482-4. Compensation between Vav-1 and Vav-2 in B cell development and antigen receptor signaling. Tedford K, Nitschke L, Girkontaite I, Charlesworth A, Chan G, Sakk V, Barbacid M, Fischer KD. Abteilung Physiologische Chemie, Universitat Ulm, Albert-Einstein-Allee 11, D-89069 Ulm, Germany. Vav-1 and Vav-2 are closely related Dbl-homology GTP exchange factors (GEFs) for Rho GTPases. Mutation of Vav-1 disrupts T cell development and T cell antigen receptor-induced activation, but has comparatively little effect on B cells. We found that combined deletion of both Vav-1 and Vav-2 in mice resulted in a marked reduction in mature B lymphocyte numbers. Vav-1(-/-)Vav-2(-/-) B cells were unresponsive to B cell antigen receptor (BCR)-driven proliferation in vitro and to thymus-independent antigen in vivo. BCR-stimulated intracellular calcium mobilization was greatly impaired in Vav-1(-/-)Vav-2(-/-) B cells. These findings establish a role for Vav-2 in BCR calcium signaling and reveal that the Vav family of GEFs is critical to B cell development and function. PMID 11376343 [PubMed - indexed for MEDLINE] Fyn-deficient mice exhibit a remarkably specific lymphoid defect thymocytes are refractile to stimulation through the TCR with mitogen or antigen. 1 Cell. 1992 Sep 4;70(5) 751-63. Defective T cell receptor signaling in mice lacking the thymic isoform of p59fyn. Appleby MW, Gross JA, Cooke MP, Levin SD, Qian X, Perlmutter RM. Howard Hughes Medical Institute, Department of Immunology, University of Washington, Seattle 98195. Considerable evidence supports the hypothesis that the nonreceptor protein tyrosine kinase p59fyn participates in signal transduction from the T cell receptor (TCR). To examine this hypothesis in detail, we have produced mice that lack the thymic isoform of p59fyn but retain expression of the brain isoform of the protein. fynTnull mice exhibit a remarkably specific lymphoid defect thymocytes are refractile to stimulation through the TCR with mitogen or antigen, while peripheral T cells, following what appears to be a normal maturation sequence, reacquire significant signaling capabilities. These data confirm that p59fynT plays a pivotal role in TCR signal transduction and demonstrate that additional developmentally regulated signaling components also contribute to TCR-induced lymphocyte activation. PMID 1516132 [PubMed - indexed for MEDLINE] Lck-deficient mice show a pronounced thymic atrophy, with a dramatic reduction in the double-positive (CD4+CD8+) thymocyte population. 1 Nature. 1992 May 14;357(6374) 161-4. Comment in Nature. 1993 Jan 21;361(6409) 213. Profound block in thymocyte development in mice lacking p56lck. Molina TJ, Kishihara K, Siderovski DP, van Ewijk W, Narendran A, Timms E, Wakeham A, Paige CJ, Hartmann KU, Veillette A, et al. Ontario Cancer Institute, University of Toronto, Canada. The protein Lck (p56lck) has a relative molecular mass of 56,000 and belongs to the Src family of tyrosine kinases. It is expressed exclusively in lymphoid cells, predominantly in thymocytes and peripheral T cells. Lck associates specifically with the cytoplasmic domains of both CD4 and CD8 T-cell surface glycoproteins and interacts with the beta-chain of the interleukin-2 receptor, which implicates Lck activity in signal transduction during thymocyte ontogeny and activation of mature T cells. Here we generate an lck null mutation by homologous recombination in embryonic stem cells to evaluate the role of p56lck in T-cell development and activation. Lck-deficient mice show a pronounced thymic atrophy, with a dramatic reduction in the double-positive (CD4+CD8+) thymocyte population. Mature, single-positive thymocytes are not detectable in these mice and there are only very few peripheral T cells. These results illustrate the crucial role of this T-cell-specific tyrosine kinase in the thymocyte development. PMID 1579166 [PubMed - indexed for MEDLINE] T cell from mice deficient in LCK is required for normal signal transduction through the TCR. 1 Cell. 1992 Aug 21;70(4) 585-93. Genetic evidence for the involvement of the lck tyrosine kinase in signal transduction through the T cell antigen receptor. Straus DB, Weiss A. Howard Hughes Medical Institute, Department of Medicine, University of California, San Francisco 94143. Signaling through the T cell antigen receptor (TCR) results both in rapid increases in tyrosine phosphorylation on a number of proteins and in the activation of the phosphatidylinositol pathway. It is not clear how stimulation of the TCR leads to these signaling events. Mutants of the Jurkat T cell line have been previously isolated that fail to show increases in calcium following receptor stimulation. Analysis of one of these mutants, JCaM1, which is defective in the induction of tyrosine phosphorylation, revealed a defect in the expression of functional lck tyrosine kinase. The lack of lck activity was caused in part by a splicing defect. Expression of the lck cDNA in JCaM1 restores the ability of the cell to respond to TCR stimulation. These results indicate that lck is required for normal signal transduction through the TCR. PMID 1505025 [PubMed - indexed for MEDLINE] T cells from mice deficient in SLAP-130/Fyb show markedly impaired proliferation. 1 Science. 2001 Sep 21;293(5538) 2263-5. Coupling of the TCR to integrin activation by Slap-130/Fyb. Peterson EJ, Woods ML, Dmowski SA, Derimanov G, Jordan MS, Wu JN, Myung PS, Liu QH, Pribila JT, Freedman BD, Shimizu Y, Koretzky GA. The Abramson Family Cancer Research Institute, Department of Medicine, School of Medicine, University of Pennsylvania, Philadelphia, PA, 19104, USA. SLAP-130/Fyb (SLP-76-associated phosphoprotein or Fyn-binding protein; also known as Fyb/Slap) is a hematopoietic-specific adapter, which associates with and modulates function of SH2-containing leukocyte phosphoprotein of 76 kilodaltons (SLP-76). T cells from mice lacking SLAP-130/Fyb show markedly impaired proliferation following CD3 engagement. In addition, the T cell receptor (TCR) in SLAP-130/Fyb mutant cells fails to enhance integrin-dependent adhesion. Although TCR-induced actin polymerization is normal, TCR-stimulated clustering of the integrin LFA-1 is defective in SLAP-130/Fyb-deficient cells. These data indicate that SLAP-130/Fyb is important for coupling TCR-mediated actin cytoskeletal rearrangement with activation of integrin function, and for T cells to respond fully to activating signals. PMID 11567141 [PubMed - indexed for MEDLINE] B cell of chicken deficient ITK reduce IP3 generation and phospholipase C gamma 2 tyrosine phosphorylation. 1 J Exp Med. 1996 Jul 1;184(1) 31-40. A role for Bruton's tyrosine kinase in B cell antigen receptor-mediated activation of phospholipase C-gamma 2. Takata M, Kurosaki T. Department of Oncology and Immunology, Wyeth-Ayerst Research, Pearl River, New York 10965, USA. Defects in the gene encoding Bruton's tyrosine kinase (Btk) result in a disease called X-linked agammaglobulinemia, in which there is a profound decrease of mature B cells due to a block in B cell development. Recent studies have shown that Btk is tyrosine phosphorylated and activated upon B cell antigen receptor (BCR) stimulation. To elucidate the functions of this kinase, we examined BCR signaling of DT40 B cells deficient in Btk. Tyrosine phosphorylation of phospholipase C (PLC)-gamma 2 upon receptor stimulation was significantly reduced in the mutant cells, leading to the loss of both BCR-coupled phosphatidylinositol hydrolysis and calcium mobilization. Pleckstrin homology and Src-homology 2 domains of Btk were required for PLC-gamma 2 activation. Since Syk is also required for the BCR-induced PLC-gamma 2 activation, our findings indicate that PLC-gamma 2 activation is regulated by Btk and Syk through their concerted actions. PMID 8691147 [PubMed - indexed for MEDLINE] T cell of mice deficient ITK reduce IP3 generation and phospholipase C gamma 1 tyrosine phosphorylation. 1 J Exp Med. 1998 May 18;187(10) 1721-7. T cell receptor-initiated calcium release is uncoupled from capacitative calcium entry in Itk-deficient T cells. Liu KQ, Bunnell SC, Gurniak CB, Berg LJ. Program of Immunology, Division of Medical Sciences, Harvard University, Boston, Massachusetts 02115, USA. Itk, a Tec family tyrosine kinase, plays an important but as yet undefined role in T cell receptor (TCR) signaling. Here we show that T cells from Itk-deficient mice have a TCR-proximal signaling defect, resulting in defective interleukin 2 secretion. Upon TCR stimulation, Itk-/- T cells release normal amounts of calcium from intracellular stores, but fail to open plasma membrane calcium channels. Since thapsigargin-induced store depletion triggers normal calcium entry in Itk-/- T cells, an impaired biochemical link between store depletion and channel opening is unlikely to be responsible for this defect. Biochemical studies indicate that TCR-induced inositol 1,4,5 tris-phosphate (IP3) generation and phospholipase C gamma1 tyrosine phosphorylation are substantially reduced in Itk-/- T cells. In contrast, TCR-zeta and ZAP-70 are phosphorylated normally, suggesting that Itk functions downstream of, or in parallel to, ZAP-70 to facilitate TCR-induced IP3 production. These findings support a model in which quantitative differences in cytosolic IP3 trigger distinct responses, and in which only high concentrations of IP3 trigger the influx of extracellular calcium. PMID 9584150 [PubMed - indexed for MEDLINE] T cell of mice deficient ITK have failure of Th2 development. 1 Immunity. 1999 Oct;11(4) 399-409. Impaired NFATc translocation and failure of Th2 development in Itk-deficient CD4+ T cells. Fowell DJ, Shinkai K, Liao XC, Beebe AM, Coffman RL, Littman DR, Locksley RM. Department of Medicine, University of California San Francisco 94143, USA. Naive Itk-deficient CD4+ T cells were unable to establish stable IL-4 production, even when primed in Th2-inducing conditions. In contrast, IFNgamma production was little affected. Failure to express IL-4 occurred even among cells that had gone through multiple cell divisions and was associated with a delay in the kinetics and magnitude of NFATc nuclear localization. IL-4 production was restored genetically by retroviral reconstitution of Itk or biochemically by augmenting the calcium flux with ionomycin. In vivo, Itk-deficient mice were unable to establish functional Th2 cells. Development of protective Th1 cells was unimpeded. These data define a nonredundant role for Itk in modulating signals from the TCR/CD28 pathways that are specific for the establishment of stable IL-4 but not IFNgamma expression. PMID 10549622 [PubMed - indexed for MEDLINE] Mice deficient in ITK have reduced proliferative responses to MHC stimulation and to anti-TCR cross-linking 1 Immunity. 1995 Dec;3(6) 757-69. Altered T cell receptor signaling and disrupted T cell development in mice lacking Itk. Liao XC, Littman DR. Department of Microbiology and Immunology, University of California, San Francisco 94143-0414, USA. Itk is a T cell protein tyrosine kinase (PTK) that, along with Btk and Tec, belongs to a family of cytoplasmic PTKs with N-terminal pleckstrin homology domains. Btk plays a critical role in B lymphocyte development. To determine whether Itk has an analogous role in T lymphocytes, we used gene targeting to prepare mice lacking expression of Itk. Such animals had decreased numbers of mature thymocytes, an effect most clearly observed in mice expressing T cell receptor (TCR) transgenes. Mature T cells from Itk-deficient mice had reduced proliferative responses to allogeneic MHC stimulation and to anti-TCR cross-linking, but responded normally to stimulation with phorbol ester plus ionomycin or with IL-2. These results provide genetic evidence that Itk is involved in T cell development and also suggest that Itk has an important role in proximal events in TCR-mediated signaling pathways. PMID 8777721 [PubMed - indexed for MEDLINE] Mutations in Btk cause X-linked immunodeficiency. 1 Semin Immunol. 1998 Aug;10(4) 309-16. Btk function in B cell development and response. Satterthwaite AB, Li Z, Witte ON. Department of Microbiology and Molecular Genetics, University of California, Los Angeles 90095-1662, USA. Mutations in Bruton's tyrosine kinase (Btk) result in the B cell immunodeficiencies XLA in humans and Xid in mice. Both the maintenance of peripheral B cell numbers and their response to B cell antigen receptor (BCR) crosslinking depend on Btk. Btk integrates signals from multiple cell surface receptors, including BCR and G-protein coupled receptors. These Btk dependent signals control B cell proliferation and survival by mediating Ca2+ flux, activating JNK and p38 and inducing cell cycle regulatory genes. Publication Types Review PMID 9695187 [PubMed - indexed for MEDLINE] Gads(GRAP2) has a role in thymocyte proliferaction for maturation of T-cells. 1 Science. 2001 Mar 9;291(5510) 1987-91. Requirement for the SLP-76 adaptor GADS in T cell development. Yoder J, Pham C, Iizuka YM, Kanagawa O, Liu SK, McGlade J, Cheng AM. Medical Scientist Training Program, Washington University School of Medicine, St. Louis, MO 63110, USA. GADS is an adaptor protein implicated in CD3 signaling because of its ability to link SLP-76 to LAT. A GADS-deficient mouse was generated by gene targeting, and the function of GADS in T cell development and activation was examined. GADS- CD4-CD8- thymocytes exhibited a severe block in proliferation but still differentiated into mature T cells. GADS- thymocytes failed to respond to CD3 cross-linking in vivo and were impaired in positive and negative selection. Immunoprecipitation experiments revealed that the association between SLP-76 and LAT was uncoupled in GADS- thymocytes. These observations indicate that GADS is a critical adaptor for CD3 signaling. PMID 11239162 [PubMed - indexed for MEDLINE] Gads(GRAP2) has a role for homeostatic proliferaction in B cells. 1 Eur J Immunol. 2005 Apr;35(4) 1184-92. Expression and function of the adaptor protein Gads in murine B cells. Yankee TM, Draves KE, Clark EA. Department of Immunology, University of Washington, Seattle, USA. tyankee@kumc.edu Nearly all hematopoietic receptors are dependent on adaptor proteins for the activation of downstream signaling pathways. The Gads adaptor protein is expressed in many hematopoietic tissues, including bone marrow, lymph node, and spleen. Using intracellular staining, we detected Gads protein in a number cells, including B cells, T cells, NK cells, monocytes, and plasmacytoid DC, but not in macrophages, neutrophils, or monocyte-derived DC. In the B cell compartment, Gads was first expressed after immature B cells leave the bone marrow and was down-regulated after B cell antigen receptor (BCR) ligation. Female Gads(-/-) mice had increased numbers of splenic B cells, as compared to female Gads(+/+) mice, suggesting a role for Gads in B cell homeostasis. Although B cell production and turnover of splenic B cell subsets appeared normal in Gads(-/-) mice, homeostatic proliferation was significantly impaired in Gads(-/-) B cells. Whereas BCR ligation can induce apoptosis in wild-type transitional stage 1 (T1) B cells, Gads(-/-) T1 B cells were resistant to BCR-induced apoptosis. Gads(-/-) B cells also showed increased BCR-mediated calcium mobilization. We conclude that Gads may have a negative regulatory role in signaling through survival pathways, and is necessary for normal homeostatic proliferation in B cells. PMID 15761845 [PubMed - indexed for MEDLINE] Grap negatively regulates T-cell proliferation. 1 Mol Cell Biol. 2002 May;22(10) 3230-6. Grap negatively regulates T-cell receptor-elicited lymphocyte proliferation and interleukin-2 induction. Shen R, Ouyang YB, Qu CK, Alonso A, Sperzel L, Mustelin T, Kaplan MH, Feng GS. Program in Signal Transduction Research, The Burnham Institute, La Jolla, California 92037, USA. Grb-2-related adaptor protein (Grap) is a Grb2-like SH3-SH2-SH3 adaptor protein with expression restricted to lymphoid tissues. Grap(-/-) lymphocytes isolated from targeted Grap-deficient mice exhibited enhanced proliferation, interleukin-2 production, and c-fos induction in response to mitogenic T-cell receptor (TCR) stimulation, compared to wild-type cells. Ectopic expression of Grap led to a suppression of Elk-1-directed transcription induced by the Ras/Erk pathway, without having effects on gene expression mediated by Jnk and p38 mitogen-activated protein kinases. Together, these data suggest that Grap, unlike Grb2, acts as a negative regulator of TCR-stimulated intracellular signaling by downregulating signal relay through the Ras/Erk pathway. PMID 11971956 [PubMed - indexed for MEDLINE] Gab2 is a substrate of ZAP-70 and functions as a switch molecule toward inhibition of TCR signal transuduction. 1 J Biol Chem. 2001 Nov 30;276(48) 45175-83. Epub 2001 Sep 25. Docking protein Gab2 is phosphorylated by ZAP-70 and negatively regulates T cell receptor signaling by recruitment of inhibitory molecules. Yamasaki S, Nishida K, Hibi M, Sakuma M, Shiina R, Takeuchi A, Ohnishi H, Hirano T, Saito T. Molecular Genetics, Chiba University Graduate School of Medicine, Chiba 260-8670, Japan. To maintain various T cell responses and immune equilibrium, activation signals triggered by T cell antigen receptor (TCR) must be regulated by inhibitory signals. Gab2, an adaptor protein of the insulin receptor substrate-1 family, has been shown to be involved in the downstream signaling from cytokine receptors. We investigated the functional role of Gab2 in TCR-mediated signal transduction. Gab2 was phosphorylated by ZAP-70 and co-precipitated with phosphoproteins, such as ZAP-70, LAT, and CD3zeta, upon TCR stimulation. Overexpression of Gab2 in Jurkat cells or antigen-specific T cell hybridomas resulted in the inhibition of NF-AT activation, interleukin-2 production, and tyrosine phosphorylation. The structure-function relationship of Gab2 was analyzed by mutants of Gab2. The Gab2 mutants lacking SHP-2-binding sites mostly abrogated the inhibitory activity of Gab2, but its inhibitory function was restored by fusing to active SHP-2 as a chimeric protein. A mutant with defective phosphatidylinositol 3-kinase binding capacity also impaired the inhibitory activity, and the pleckstrin homology domain-deletion mutant revealed a crucial function of the pleckstrin homology domain for localization to the plasma membrane. These results suggest that Gab2 is a substrate of ZAP-70 and functions as a switch molecule toward inhibition of TCR signal transduction by mediating the recruitment of inhibitory molecules to the TCR signaling complex. PMID 11572860 [PubMed - indexed for MEDLINE] B cell signaling causes tyrosine phosphorylation of Gab1, and in turn SHP2 bind to Gab1 Gab1 phosophorylation potentiate the phosphorylation of Akt, PI3K-dependent response. 1 J Biol Chem. 2001 Apr 13;276(15) 12257-65. Epub 2001 Jan 22. The Gab1 docking protein links the b cell antigen receptor to the phosphatidylinositol 3-kinase/Akt signaling pathway and to the SHP2 tyrosine phosphatase. Ingham RJ, Santos L, Dang-Lawson M, Holgado-Madruga M, Dudek P, Maroun CR, Wong AJ, Matsuuchi L, Gold MR. Departments of Microbiology and Immunology and Zoology, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada. B cell antigen receptor (BCR) signaling causes tyrosine phosphorylation of the Gab1 docking protein. This allows phosphatidylinositol 3-kinase (PI3K) and the SHP2 tyrosine phosphatase to bind to Gab1. In this report, we tested the hypothesis that Gab1 acts as an amplifier of PI3K- and SHP2-dependent signaling in B lymphocytes. By overexpressing Gab1 in the WEHI-231 B cell line, we found that Gab1 can potentiate BCR-induced phosphorylation of Akt, a PI3K-dependent response. Gab1 expression also increased BCR-induced tyrosine phosphorylation of SHP2 as well as the binding of Grb2 to SHP2. We show that the pleckstrin homology (PH) domain of Gab1 is required for BCR-induced phosphorylation of Gab1 and for Gab1 participation in BCR signaling. Moreover, using confocal microscopy, we show that BCR ligation can induce the translocation of Gab1 from the cytosol to the plasma membrane and that this requires the Gab1 PH domain as well as PI3K activity. These findings are consistent with a model in which the binding of the Gab1 PH domain to PI3K-derived lipids brings Gab1 to the plasma membrane, where it can be tyrosine-phosphorylated and then act as an amplifier of BCR signaling. PMID 11278704 [PubMed - indexed for MEDLINE] RasGRP1 mediates Ras activation following TCR stimulatioin. RasGRP1 and RasGRP3 induces RAS activation in B-cell to response to T-cell stimulation. 1 Immunol Lett. 2006 May 15;105(1) 77-82. Epub 2006 Feb 20. The role of RasGRPs in regulation of lymphocyte proliferation. Coughlin JJ, Stang SL, Dower NA, Stone JC. Department of Biochemistry, University of Alberta, Edmonton, Alta., Canada T6G 2H7. RasGRP1 links TCR signaling to Ras in T cells, while both RasGRP1 and RasGRP3 link BCR signaling to Ras in B cells. T cells deficient in RasGRP1 have defective proliferative responses as do B cells deficient in both RasGRP1 and RasGRP3, confirming the importance of Ras activation in lymphocyte proliferation. While aged Rasgrp1-/- mice develop late-onset autoimmunity characterized by splenomegaly and the presence of anti-nuclear antibodies (ANA), the additional loss of RasGRP3 expression inhibits this phenotype. We show here that the autoimmunity in Rasgrp1-/- mice is T cell dependent. Compared to wildtype, Rasgrp1-/- T cells induce greater in vitro B cell proliferation that is due, at least in part, to increased production of interleukin-4 (IL-4). Rasgrp1 Rasgrp3 double mutant B cells are less responsive to this T cell stimulation. The reduced double mutant B cell proliferative response was paralleled by decreased induction of cyclin D2 upon stimulation with IL-4 and anti-IgM. Taken together these results suggest that double mutant mice fail to generate autoimmunity due to their decreased B cell cyclin D2 accumulation, and thus proliferation, in response to the elevated levels of IL-4 produced by mutant T cells. PMID 16530850 [PubMed - indexed for MEDLINE] Grb2-hSos1-PLCgamma1-p36/p38-ZAP70 complexes localize in the vicinity of TCR-zeta 1 J Biol Chem. 1995 Aug 4;270(31) 18428-36. Ligation of the T-cell antigen receptor (TCR) induces association of hSos1, ZAP-70, phospholipase C-gamma 1, and other phosphoproteins with Grb2 and the zeta-chain of the TCR. Nel AE, Gupta S, Lee L, Ledbetter JA, Kanner SB. Department of Medicine, UCLA School of Medicine 90024, USA. Signaling by the T-cell antigen receptor (TCR) involves both phospholipase C (PLC)-gamma 1 and p21ras activation. While failing to induce Shc/Grb2 association, ligation of the TCR/CD3 receptor in Jurkat T-cells induced hSos1-Grb2 complexes. In addition to hSos1, Grb2 participates in the formation of a tyrosine phosphoprotein complex that includes 145-, 95-, 70-, 54-, and 36-38-kDa proteins. p145 was identified as PLC-gamma 1 and p70 as the protein tyrosine kinase, ZAP-70. Although of the same molecular weight, p95 was not recognized by an anti-serum to p95 Vav. The SH2 domains of Grb2 and PLC-gamma 1 were required for the formation of this protein complex. In anti-CD3-treated cells, Grb2 redistributed from the cytosol to a particulate cell compartment along with p36/p38, ZAP-70, and PLC-gamma 1. Part of the Grb2 complex associated with the particulate compartment could be extracted with Nonidet P-40, while the rest was Nonidet P-40 insoluble. In both the detergent-soluble and -insoluble fractions, Grb2 coimmunoprecipitated with the zeta-chain of the TCR. Taken together, these results indicate that anti-CD3 induces Grb2-hSos1-PLC-gamma 1-p36/p38-ZAP70 complexes, which localize in the vicinity of TCR-zeta. PMID 7629168 [PubMed - indexed for MEDLINE] Gads(Grap2) plays an important role in T-cell signaling via its association with SLP-76 and LAT. 1 Curr Biol. 1999 Jan 28;9(2) 67-75. The hematopoietic-specific adaptor protein gads functions in T-cell signaling via interactions with the SLP-76 and LAT adaptors. Liu SK, Fang N, Koretzky GA, McGlade CJ. Department of Medical Biophysics, University of Toronto, The Arthur and Sonia Labatt Brain Tumour Research Centre, Hospital for Sick Children, Research Institute, 555 University Ave, Toronto, Ontario M5G 1X8, Canada. BACKGROUND The adaptor protein Gads is a Grb2-related protein originally identified on the basis of its interaction with the tyrosine-phosphorylated form of the docking protein Shc. Gads protein expression is restricted to hematopoietic tissues and cell lines. Gads contains a Src homology 2 (SH2) domain, which has previously been shown to have a similar binding specificity to that of Grb2. Gads also possesses two SH3 domains, but these have a distinct binding specificity to those of Grb2, as Gads does not bind to known Grb2 SH3 domain targets. Here, we investigated whether Gads is involved in T-cell signaling. RESULTS We found that Gads is highly expressed in T cells and that the SLP-76 adaptor protein is a major Gads-associated protein in vivo. The constitutive interaction between Gads and SLP-76 was mediated by the carboxy-terminal SH3 domain of Gads and a 20 amino-acid proline-rich region in SLP-76. Gads also coimmunoprecipitated the tyrosine-phosphorylated form of the linker for activated T cells (LAT) adaptor protein following cross-linking of the T-cell receptor; this interaction was mediated by the Gads SH2 domain. Overexpression of Gads and SLP-76 resulted in a synergistic augmentation of T-cell signaling, as measured by activation of nuclear factor of activated T cells (NFAT), and this cooperation required a functional Gads SH2 domain. CONCLUSIONS These results demonstrate that Gads plays an important role in T-cell signaling via its association with SLP-76 and LAT. Gads may promote cross-talk between the LAT and SLP-76 signaling complexes, thereby coupling membrane-proximal events to downstream signaling pathways. PMID 10021361 [PubMed - indexed for MEDLINE] Lck is required for normal signal transduction through the TCR. 1 Cell. 1992 Aug 21;70(4) 585-93. Genetic evidence for the involvement of the lck tyrosine kinase in signal transduction through the T cell antigen receptor. Straus DB, Weiss A. Howard Hughes Medical Institute, Department of Medicine, University of California, San Francisco 94143. Signaling through the T cell antigen receptor (TCR) results both in rapid increases in tyrosine phosphorylation on a number of proteins and in the activation of the phosphatidylinositol pathway. It is not clear how stimulation of the TCR leads to these signaling events. Mutants of the Jurkat T cell line have been previously isolated that fail to show increases in calcium following receptor stimulation. Analysis of one of these mutants, JCaM1, which is defective in the induction of tyrosine phosphorylation, revealed a defect in the expression of functional lck tyrosine kinase. The lack of lck activity was caused in part by a splicing defect. Expression of the lck cDNA in JCaM1 restores the ability of the cell to respond to TCR stimulation. These results indicate that lck is required for normal signal transduction through the TCR. PMID 1505025 [PubMed - indexed for MEDLINE] ZAP-70 plays crucial roles in T-cell activation and development. Syk triggers cellular activation in T-cell. 1 Mol Cell Biol. 1998 Mar;18(3) 1388-99. Genetic evidence for differential coupling of Syk family kinases to the T-cell receptor reconstitution studies in a ZAP-70-deficient Jurkat T-cell line. Williams BL, Schreiber KL, Zhang W, Wange RL, Samelson LE, Leibson PJ, Abraham RT. Department of Immunology, Mayo Clinic, Rochester, Minnesota 55905, USA. T-cell antigen receptor (TCR) engagement activates multiple protein tyrosine kinases (PTKs), including the Src family member, Lck, and the Syk-related PTK, ZAP-70. Studies in ZAP-70-deficient humans have demonstrated that ZAP-70 plays crucial roles in T-cell activation and development. However, progress toward a detailed understanding of the regulation and function of ZAP-70 during TCR signaling has been hampered by the lack of a suitable T-cell model for biochemical and genetic analyses. In this report, we describe the isolation and phenotypic characterization of a Syk- and ZAP-70-negative somatic mutant derived from the Jurkat T-cell line. The P116 cell line displays severe defects in TCR-induced signaling functions, including protein tyrosine phosphorylation, intracellular Ca2+ mobilization, and interleukin-2 promoter-driven transcription. These signaling defects were fully reversed by reintroduction of catalytically active versions of either Syk or ZAP-70 into the P116 cells. However, in contrast to ZAP-70 expression, Syk expression triggered a significant degree of cellular activation in the absence of TCR ligation. Transfection experiments with ZAP-70-Syk chimeric proteins indicated that both the amino-terminal regulatory regions and the carboxy-terminal catalytic domains of Syk and ZAP-70 contribute to the distinctive functional properties of these PTKs. These studies underscore the crucial role of ZAP-70 in TCR signaling and offer a powerful genetic model for further analyses of ZAP-70 regulation and function in T cells. PMID 9488454 [PubMed - indexed for MEDLINE]