isolation-of-cancer-stem-cells-from-human-prostate-cancer-samples

Isolation of Cancer Stem Cells From Human Prostate Cancer Samples

The cancer stem cell (CSC) model has been considerably revisited over the last two decades. During this time CSCs have been identified and directly ...

generation-prostate-cancer-patient-derived-xenograft-models-from

Generation of Prostate Cancer Patient Derived Xenograft Models from Circulating Tumor Cells

Patient derived xenograft (PDX) models are gaining popularity in cancer research and are used for preclinical drug evaluation, biomarker identification, ...

generation-prostate-cancer-cell-models-resistance-to-anti-mitotic

Generation of Prostate Cancer Cell Models of Resistance to the Anti-mitotic Agent Docetaxel

Microtubule targeting agents (MTAs) are a mainstay in the treatment of a wide range of tumors. However, acquired resistance to chemotherapeutic drugs is a ...

isolation-characterization-tumor-initiating-cells-from-sarcoma

Isolation and Characterization of Tumor-initiating Cells from Sarcoma Patient-derived Xenografts

The existence and importance of tumor-initiating cells (TICs) have been supported by increasing evidence during the past decade. These TICs have been ...

Memorial Sloan-Kettering Cancer Center

Icahn School of Medicine at Mount Sinai

When DNA synthesis is inhibited, DNA replication checkpoint is activated to prevent mitosis entry without fully replicated DNA. In Xenopus, caffeine-sensitive kinases [ataxia telangiectasia mutated (ATM) and ATM-related protein (ATR)] are essential in this checkpoint response, but in mammal cells an ATR/ATM-independent checkpoint response to DNA synthesis inhibition exists. Using HeLa cells, which have a caffeine-insensitive checkpoint response, we have analyzed here which molecules known to be involved in the DNA replication checkpoint participate in the caffeine-insensitive response. When DNA synthesis was inhibited in the presence of UCN01 or after knocking down Chk1 expression [Chk1 small interfering RNA (siRNA)], HeLa cells entered into aberrant mitosis. Consequently, Chk1 is essential for both the ATR/ATM-dependent and ATR/ATM-independent checkpoint response in HeLa cells. Neither wortmannin, Ly294002, nor SB202190 abrogated the caffeine-insensitive checkpoint response, indicating that DNA-PK and p38 alpha,beta are not involved in the ATR/ATM-independent Chk1 activation upon DNA synthesis inhibition. Using siRNA to knock down Rad17 and claspin, two molecules involved in sensing stalled replication forks, we also showed that claspin but not Rad17 is essential for the ATR/ATM-independent checkpoint response. Inhibition of DNA synthesis in HeLa cells led to a decrease in cyclin B1 protein accumulation that was abrogated when UCN01 was added or when claspin was knocked down. We conclude that upon DNA synthesis inhibition, Chk1 can be activated in a claspin-dependent manner independently of ATR and ATM, leading to cyclin B1 down-regulation and providing the cells of an additional mechanism to inhibit mitosis entry.

Chk1- and claspin-dependent but ATR/ATM- and Rad17-independent DNA replication checkpoint response in HeLa cells.

Cell cycle checkpoint abrogation, especially the inhibition of Chk1 in combination with DNA-damaging treatments, has been proposed as a promising way of sensitizing cancer cells. However, less is known about the possibility to selectively affect tumor cells when they are treated with agents that block DNA synthesis in combination with replication checkpoint inhibitors. Here, we present clear insights in the different responses of tumor and non-transformed cells to the inhibition of DNA replication with hydroxyurea in combination with checkpoint abrogation via inhibition of Ataxia telangiectasia-mutated- (ATM) and Rad3-related/ATM (ATR/ATM) and Chk1 kinases. Interestingly, we find that non-transformed cell lines activate ATR/ATM- and Chk1-independent pathways in response to replication inhibition to prevent mitotic entry with unreplicated DNA. In contrast, tumor cell lines such as HCT116 and HeLa cells rely entirely on Chk1 activity for a proper response to replication inhibitors. Our results show that p38 is activated in response to hydroxyurea treatment and collaborates with Chk1 to prevent mitotic entry in non-transformed cell lines by maintaining cyclin B1/Cdk1 complexes inactive. Furthermore, DNA replication arrest down-regulates cyclin B1 promoter activity in non-transformed cells, but not in tumor cells in a Chk1- and p38-independent way. Thus, our data show that non-transformed cells present a more robust DNA replication checkpoint response compared with tumor cells that involves activation of the p38 pathway. We show that some of these responses to replication block can be lost in tumor cells, causing a defective checkpoint and providing a rationale for tumor-selective effects of combined therapies.

Different S/M checkpoint responses of tumor and non tumor cell lines to DNA replication inhibition.

&#x51B3;&#x5B9A;&#x8FDB;&#x5165;&#x6709;&#x4E1D;&#x5206;&#x88C2;&#x88AB;&#x4ECB;&#x5BFC;&#x86CB;&#x767D;&#x8C03;&#x63A7;&#x7EC6;&#x80DE;&#x5468;&#x671F;&#x86CB;&#x767D; B Cdk1 &#x590D;&#x6742;&#x7684;&#x6FC0;&#x6D3B;&#x7684;&#x7F51;&#x7EDC;&#x3002;&#x5728;&#x8FD9;&#x4E2A;&#x7F51;&#x7EDC;&#x5185;&#x51E0;&#x4E2A;&#x79EF;&#x6781;&#x7684;&#x53CD;&#x9988;&#x56DE;&#x8DEF;&#x53EF;&#x4EE5;&#x653E;&#x5927;&#x7EC6;&#x80DE;&#x5468;&#x671F;&#x86CB;&#x767D; B Cdk1 &#x6FC0;&#x6D3B;&#xFF0C;&#x4EE5;&#x4FDD;&#x8BC1;&#x5B8C;&#x5168;&#x81F4;&#x529B;&#x4E8E;&#x6709;&#x4E1D;&#x5206;&#x88C2;&#x72B6;&#x6001;&#xFF0C;&#x4E00;&#x65E6;&#x53D6;&#x5F97;&#x4E86;&#x8FDB;&#x5165;&#x6709;&#x4E1D;&#x5206;&#x88C2;&#x7684;&#x51B3;&#x5B9A;&#x3002;&#x7136;&#x800C;&#xFF0C;&#x8BC1;&#x636E;&#x7D2F;&#x79EF;&#x53CD;&#x9988;&#x5FAA;&#x73AF;&#x7684;&#x51E0;&#x4E2A;&#x7EC4;&#x4EF6;&#x5197;&#x4F59;&#x7684;&#x6B63;&#x5E38;&#x7684;&#x7EC6;&#x80DE;&#x5206;&#x88C2;&#x8FC7;&#x7A0B;&#x4E2D; B Cdk1 &#x6D3B;&#x5316;&#x7EC6;&#x80DE;&#x5468;&#x671F;&#x3002;&#x5C3D;&#x7BA1;&#x5982;&#x6B64;&#xFF0C;&#x5B9A;&#x4E49;&#x7684;&#x53CD;&#x9988;&#x5FAA;&#x73AF;&#x6210;&#x4E3A;&#x5FC5;&#x987B;&#x4FC3;&#x8FDB;&#x6709;&#x4E1D;&#x5206;&#x88C2;&#x7684;&#x6761;&#x76EE;&#xFF0C;&#x5F53;&#x6B63;&#x5E38;&#x7EC6;&#x80DE;&#x5468;&#x671F;&#x8FDB;&#x7A0B;&#x6444;&#x52A8;&#x3002;&#x6700;&#x8FD1;&#x7684;&#x6570;&#x636E;&#x8868;&#x660E;&#x81F3;&#x5C11;&#x4E09;&#x4E2A; Plk1 &#x76F8;&#x5173;&#x53CD;&#x9988;&#x5FAA;&#x73AF;&#x5B58;&#x5728;&#xFF0C;&#x52A0;&#x5F3A;&#x4E0D;&#x540C;&#x7EA7;&#x522B;&#x7684;&#x7EC6;&#x80DE;&#x5468;&#x671F;&#x86CB;&#x767D; B Cdk1 &#x6FC0;&#x6D3B;&#x3002;&#x8FD9;&#x9879;&#x5BA1;&#x67E5;&#xFF0C;&#x5728;&#x6211;&#x4EEC;&#x8BA8;&#x8BBA;&#x5404;&#x79CD;&#x8C03;&#x8282;&#x7EC6;&#x80DE;&#x5468;&#x671F;&#x86CB;&#x767D;&#x5728;&#x4E0D;&#x540C;&#x6761;&#x4EF6;&#x4E0B;&#xFF0C;B Cdk1 &#x6FC0;&#x6D3B;&#xFF0C;&#x5176;&#x6FC0;&#x6D3B;&#x7684;&#x65F6;&#x95F4;&#x548C;&#x96BE;&#x4EE5;&#x6349;&#x6478;&#x7684;&#x89E6;&#x53D1;&#x5668;&#xFF0C;&#x63A7;&#x5236;&#x7EC6;&#x80DE;&#x7684;&#x6709;&#x4E1D;&#x5206;&#x88C2;&#x8FDB;&#x5165;&#x7684;&#x53EF;&#x80FD;&#x8EAB;&#x4EFD;&#x7684;&#x53CD;&#x9988;&#x5FAA;&#x73AF;&#x7684;&#x4F5C;&#x7528;&#x3002;

&#x8F93;&#x5165;&#x6709;&#x4E1D;&#x5206;&#x88C2;&#x7684;&#x51B3;&#x5B9A;&#xFF1A; &#x53CD;&#x9988;&#x548C;&#x6709;&#x4E1D;&#x5206;&#x88C2;&#x8FDB;&#x5165;&#x7F51;&#x7EDC;&#x4E2D;&#x7684;&#x5197;&#x4F59;&#x3002;

The decision to enter mitosis is mediated by a network of proteins that regulate activation of the cyclin B-Cdk1 complex. Within this network, several positive feedback loops can amplify cyclin B-Cdk1 activation to ensure complete commitment to a mitotic state once the decision to enter mitosis has been made. However, evidence is accumulating that several components of the feedback loops are redundant for cyclin B-Cdk1 activation during normal cell division. Nonetheless, defined feedback loops become essential to promote mitotic entry when normal cell cycle progression is perturbed. Recent data has demonstrated that at least three Plk1-dependent feedback loops exist that enhance cyclin B-Cdk1 activation at different levels. In this review, we discuss the role of various feedback loops that regulate cyclin B-Cdk1 activation under different conditions, the timing of their activation, and the possible identity of the elusive trigger that controls mitotic entry in human cells.

The decision to enter mitosis: feedback and redundancy in the mitotic entry network.

La d&#xE9;cision d&#x26;#39;entrer dans la mitose est m&#xE9;di&#xE9;e par un r&#xE9;seau de prot&#xE9;ines qui r&#xE9;gulent l&#x26;#39;activation de la cycline B-Cdk1 complexe. Dans ce r&#xE9;seau, plusieurs boucles de r&#xE9;troaction positive peut amplifier cycline B-Cdk1 activation pour garantir l&#x26;#39;engagement complet dans un &#xE9;tat mitotique une fois que la d&#xE9;cision d&#x26;#39;entrer dans la mitose a &#xE9;t&#xE9; faite. Toutefois, les preuves s&#x26;#39;accumulent que plusieurs composantes des boucles de r&#xE9;troaction sont redondantes pour la cycline B-Cdk1 activation pendant la division cellulaire normale. N&#xE9;anmoins, des boucles de r&#xE9;troaction d&#xE9;finies devenu indispensable de promouvoir l&#x26;#39;entr&#xE9;e en mitose lorsque normale progression du cycle cellulaire est perturb&#xE9;e. Des donn&#xE9;es r&#xE9;centes ont montr&#xE9; qu&#x26;#39;au moins trois boucles de r&#xE9;troaction Plk1-d&#xE9;pendants existent qui am&#xE9;liorent la cycline B-Cdk1 activation &#xE0; diff&#xE9;rents niveaux. Dans cette revue, nous discutons le r&#xF4;le des boucles de r&#xE9;troaction qui r&#xE9;gulent divers cycline B-Cdk1 activation dans des conditions diff&#xE9;rentes, le moment de leur activation, et l&#x26;#39;identit&#xE9; possible de la g&#xE2;chette insaisissable qui contr&#xF4;le l&#x26;#39;entr&#xE9;e en mitose dans les cellules humaines.

La d&#xE9;cision d&#x26;#39;entrer Mitose: Commentaires et redondance dans le r&#xE9;seau l&#x26;#39;entr&#xE9;e en mitose

Die Entscheidung zur Mitose geben ist durch ein Netz von Proteine vermittelt, die Aktivierung der cyclin B-Cdk1-Komplex zu regulieren. In diesem Verbund k&ouml;nnen mehrere positive Feedback-Schleifen verst&auml;rken cyclin B-Cdk1-Aktivierung um komplette Engagement f&uuml;r eine mitotische Staat zu gew&auml;hrleisten, wenn die Entscheidung zur Mitose Eingabe get&auml;tigt wurde. Jedoch ist Beweis ansammeln, dass mehrere Komponenten der Feedback-Schleifen f&uuml;r cyclin B-Cdk1 Aktivierung w&auml;hrend der normalen Zellteilung redundant sind. Dennoch werden definierte R&uuml;ckkopplungsschleifen wesentliche F&ouml;rderung mitotische Eintrag beim normalen Zellzyklusprogression gest&ouml;rt ist. J&uuml;ngste Daten hat gezeigt, dass mindestens drei Plk1-abh&auml;ngige-Feedback-Schleifen vorhanden sind, die cyclin B-Cdk1-Aktivierung auf verschiedenen Ebenen zu verbessern. In diesem Bericht diskutieren wir die Rolle der verschiedenen Feedback-Schleifen, die Regeln von cyclin B-Cdk1-Aktivierung unter verschiedenen Bedingungen, das Timing von deren Aktivierung und die m&ouml;gliche Identit&auml;t des fl&uuml;chtigen Triggers, die mitotische Eintrag in den menschlichen Zellen steuert.

Die Entscheidung zur Mitose eingeben: Feedback und Redundanz in der mitotischen Eintrag-Netzwerk.

La decisione di entrare in mitosi &egrave; mediata da una rete di proteine che regolano l&#39;attivazione del complesso cyclin B-Cdk1. All&#39;interno di questa rete, diversi cicli di retroazione positiva possono amplificare cyclin B-Cdk1 attivazione per garantire totale impegno per uno stato mitotico, una volta che &egrave; stata presa la decisione di entrare in mitosi. Tuttavia, prove sta accumulando che diversi componenti di feedback loop sono ridondanti per ciclina B-Cdk1 attivazione durante la divisione cellulare normale. Tuttavia, definito feedback loop diventano essenziali per promuovere la voce mitotica quando la progressione del ciclo cellulare normale &egrave; perturbato. Dati recenti hanno dimostrato che almeno tre cicli di retroazione Plk1-dipendente esistano che migliorano cyclin B-Cdk1 attivazione a diversi livelli. In questa recensione, si discute il ruolo dei vari cicli di retroazione che regolano cyclin B-Cdk1 attivazione in condizioni diverse, i tempi della loro attivazione e la possibile identit&agrave; del trigger sfuggente che controlla la voce mitosi nelle cellule umane.

La decisione di entrare in mitosi: feedback e ridondanza della rete voce mitotica.

&#x6709;&#x7CF8;&#x5206;&#x88C2;&#x3092;&#x5165;&#x529B;&#x3059;&#x308B;&#x304B;&#x3069;&#x3046;&#x304B;&#x306E;&#x6C7A;&#x5B9A;&#x306F;&#x3001;&#x30B5;&#x30A4;&#x30AF;&#x30EA;&#x30F3;B-CDK1&#x8907;&#x5408;&#x4F53;&#x306E;&#x6D3B;&#x6027;&#x5316;&#x3092;&#x8ABF;&#x7BC0;&#x3059;&#x308B;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x306E;&#x30CD;&#x30C3;&#x30C8;&#x30EF;&#x30FC;&#x30AF;&#x306B;&#x3088;&#x3063;&#x3066;&#x5A92;&#x4ECB;&#x3055;&#x308C;&#x308B;&#x3002;&#x3053;&#x306E;&#x30CD;&#x30C3;&#x30C8;&#x30EF;&#x30FC;&#x30AF;&#x5185;&#x3067;&#x3001;&#x3044;&#x304F;&#x3064;&#x304B;&#x306E;&#x6B63;&#x306E;&#x30D5;&#x30A3;&#x30FC;&#x30C9;&#x30D0;&#x30C3;&#x30AF;&#x30EB;&#x30FC;&#x30D7;&#x306F;&#x3001;&#x6709;&#x7CF8;&#x5206;&#x88C2;&#x3092;&#x5165;&#x529B;&#x3059;&#x308B;&#x305F;&#x3081;&#x306E;&#x6C7A;&#x5B9A;&#x304C;&#x306A;&#x3055;&#x308C;&#x305F;&#x5F8C;&#x5206;&#x88C2;&#x72B6;&#x614B;&#x3078;&#x306E;&#x5B8C;&#x5168;&#x306A;&#x30B3;&#x30DF;&#x30C3;&#x30C8;&#x30E1;&#x30F3;&#x30C8;&#x3092;&#x78BA;&#x4FDD;&#x3059;&#x308B;&#x305F;&#x3081;&#x3001;&#x30B5;&#x30A4;&#x30AF;&#x30EA;&#x30F3;B-CDK1&#x306E;&#x6D3B;&#x6027;&#x5316;&#x3092;&#x5897;&#x5E45;&#x3059;&#x308B;&#x3053;&#x3068;&#x304C;&#x3067;&#x304D;&#x307E;&#x3059;&#x3002;&#x3057;&#x304B;&#x3057;&#x3001;&#x8A3C;&#x62E0;&#x306F;&#x3001;&#x30D5;&#x30A3;&#x30FC;&#x30C9;&#x30D0;&#x30C3;&#x30AF;&#x30EB;&#x30FC;&#x30D7;&#x306E;&#x3044;&#x304F;&#x3064;&#x304B;&#x306E;&#x30B3;&#x30F3;&#x30DD;&#x30FC;&#x30CD;&#x30F3;&#x30C8;&#x306F;&#x3001;&#x901A;&#x5E38;&#x306E;&#x7D30;&#x80DE;&#x5206;&#x88C2;&#x306E;&#x969B;&#x306B;&#x30B5;&#x30A4;&#x30AF;&#x30EA;&#x30F3;B-CDK1&#x306E;&#x6D3B;&#x6027;&#x5316;&#x306E;&#x305F;&#x3081;&#x306B;&#x5197;&#x9577;&#x3067;&#x3042;&#x308B;&#x3053;&#x3068;&#x304C;&#x84C4;&#x7A4D;&#x3055;&#x308C;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x305D;&#x308C;&#x306B;&#x3082;&#x304B;&#x304B;&#x308F;&#x3089;&#x305A;&#x3001;&#x5B9A;&#x7FA9;&#x3055;&#x308C;&#x305F;&#x30D5;&#x30A3;&#x30FC;&#x30C9;&#x30D0;&#x30C3;&#x30AF;&#x30EB;&#x30FC;&#x30D7;&#x306F;&#x3001;&#x901A;&#x5E38;&#x306E;&#x7D30;&#x80DE;&#x5468;&#x671F;&#x306E;&#x9032;&#x884C;&#x3092;&#x6442;&#x52D5;&#x3055;&#x308C;&#x305F;&#x3068;&#x304D;&#x306B;&#x5206;&#x88C2;&#x30A8;&#x30F3;&#x30C8;&#x30EA;&#x3092;&#x4FC3;&#x9032;&#x3059;&#x308B;&#x305F;&#x3081;&#x306B;&#x4E0D;&#x53EF;&#x6B20;&#x3068;&#x306A;&#x308B;&#x3002;&#x6700;&#x8FD1;&#x306E;&#x30C7;&#x30FC;&#x30BF;&#x306F;&#x3001;&#x5C11;&#x306A;&#x304F;&#x3068;&#x3082;3&#x3064;&#x306E;PLK1&#x4F9D;&#x5B58;&#x30D5;&#x30A3;&#x30FC;&#x30C9;&#x30D0;&#x30C3;&#x30AF;&#x30EB;&#x30FC;&#x30D7;&#x306F;&#x3001;&#x3055;&#x307E;&#x3056;&#x307E;&#x306A;&#x30EC;&#x30D9;&#x30EB;&#x3067;&#x30B5;&#x30A4;&#x30AF;&#x30EA;&#x30F3;B-CDK1&#x306E;&#x6D3B;&#x6027;&#x5316;&#x3092;&#x9AD8;&#x3081;&#x308B;&#x3053;&#x3068;&#x304C;&#x5B58;&#x5728;&#x3059;&#x308B;&#x3053;&#x3068;&#x3092;&#x5B9F;&#x8A3C;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x3053;&#x306E;&#x30EC;&#x30D3;&#x30E5;&#x30FC;&#x3067;&#x306F;&#x3001;&#x30B5;&#x30A4;&#x30AF;&#x30EA;&#x30F3;B-CDK1&#x7570;&#x306A;&#x308B;&#x6761;&#x4EF6;&#x4E0B;&#x3067;&#x306E;&#x6D3B;&#x6027;&#x5316;&#x3001;&#x305D;&#x308C;&#x3089;&#x306E;&#x6D3B;&#x6027;&#x5316;&#x306E;&#x30BF;&#x30A4;&#x30DF;&#x30F3;&#x30B0;&#x3001;&#x304A;&#x3088;&#x3073;&#x30D2;&#x30C8;&#x306E;&#x7D30;&#x80DE;&#x3067;&#x6709;&#x7CF8;&#x5206;&#x88C2;&#x306E;&#x30A8;&#x30F3;&#x30C8;&#x30EA;&#x3092;&#x5236;&#x5FA1;&#x3059;&#x308B;&#x3068;&#x3089;&#x3048;&#x3069;&#x3053;&#x308D;&#x306E;&#x306A;&#x3044;&#x30C8;&#x30EA;&#x30AC;&#x30FC;&#x306E;&#x53EF;&#x80FD;&#x6027;&#x306E;&#x30A2;&#x30A4;&#x30C7;&#x30F3;&#x30C6;&#x30A3;&#x30C6;&#x30A3;&#x3092;&#x8ABF;&#x7BC0;&#x3059;&#x308B;&#x69D8;&#x3005;&#x306A;&#x30D5;&#x30A3;&#x30FC;&#x30C9;&#x30D0;&#x30C3;&#x30AF;&#x30EB;&#x30FC;&#x30D7;&#x306E;&#x5F79;&#x5272;&#x3092;&#x8B70;&#x8AD6;&#x3059;&#x308B;&#x3002;

&#x6709;&#x7CF8;&#x5206;&#x88C2;&#x3092;&#x5165;&#x529B;&#x3059;&#x308B;&#x305F;&#x3081;&#x306E;&#x6C7A;&#x5B9A;&#xFF1A;&#x5206;&#x88C2;&#x30A8;&#x30F3;&#x30C8;&#x30EA;&#x30CD;&#x30C3;&#x30C8;&#x30EF;&#x30FC;&#x30AF;&#x306E;&#x30D5;&#x30A3;&#x30FC;&#x30C9;&#x30D0;&#x30C3;&#x30AF;&#x3068;&#x5197;&#x9577;&#x6027;

&#xC720;&#xC0AC; &#xBD84;&#xC5F4;&#xC744; &#xC785;&#xB825; &#xD558;&#xB3C4;&#xB85D; &#xACB0;&#xC815; cyclin B Cdk1 &#xB2E8;&#xC9C0;&#xC758; &#xD65C;&#xC131;&#xD654;&#xB97C; &#xC870;&#xC808; &#xD558;&#xB294; &#xB2E8;&#xBC31;&#xC9C8;&#xC758; &#xB124;&#xD2B8;&#xC6CC;&#xD06C;&#xC5D0; &#xC758;&#xD574; &#xC911;&#xC7AC;&#xC785;&#xB2C8;&#xB2E4;. &#xC774; &#xB124;&#xD2B8;&#xC6CC;&#xD06C; &#xB0B4;&#xC5D0;&#xC11C; &#xC5EC;&#xB7EC; &#xAC00;&#xC9C0; &#xAE0D;&#xC815;&#xC801;&#xC778; &#xD53C;&#xB4DC;&#xBC31; &#xB8E8;&#xD504; &#xC785;&#xB825; &#xC720;&#xC0AC; &#xBD84;&#xC5F4;&#xC744; &#xACB0;&#xC815; &#xD6C4; mitotic &#xC0C1;&#xD0DC;&#xB85C; &#xC644;&#xC804; &#xD55C; &#xD5CC;&#xC2E0;&#xC744; &#xBCF4;&#xC7A5; &#xD558;&#xAE30; &#xC704;&#xD574; cyclin B Cdk1 &#xD65C;&#xC131;&#xD654;&#xB97C; &#xC99D;&#xD3ED; &#xC218; &#xC788;&#xC2B5;&#xB2C8;&#xB2E4;. &#xADF8;&#xB7EC;&#xB098;, &#xC99D;&#xAC70; &#xD53C;&#xB4DC;&#xBC31; &#xB8E8;&#xD504;&#xC758; &#xC77C;&#xBD80; &#xAD6C;&#xC131; &#xC694;&#xC18C;&#xB294; &#xC911;&#xBCF5; &#xF06C; &#xC815;&#xC0C1;&#xC801;&#xC778; &#xC138;&#xD3EC; &#xBD84;&#xC5F4; &#xC911;&#xC5D0; B Cdk1 &#xD65C;&#xC131;&#xD654;&#xC5D0; &#xB300; &#xD55C; &#xCD95;&#xC801; &#xC774;&#xB2E4;. &#xADF8;&#xB7FC;&#xC5D0;&#xB3C4; &#xBD88;&#xAD6C; &#xD558; &#xACE0;, &#xC815;&#xC758; &#xB41C; &#xD53C;&#xB4DC;&#xBC31; &#xB8E8;&#xD504; &#xB420; &#xB54C; &#xC815;&#xC0C1;&#xC801;&#xC778; &#xC138;&#xD3EC; &#xC8FC;&#xAE30; &#xC9C4;&#xD589; &#xBD88;&#xC548;&#xC815; mitotic &#xD56D;&#xBAA9;&#xC744; &#xD64D;&#xBCF4; &#xD544;&#xC218;. &#xCD5C;&#xADFC; &#xB370;&#xC774;&#xD130;&#xB294; &#xC801;&#xC5B4;&#xB3C4; 3 &#xAC1C;&#xC758; Plk1 &#xC885;&#xC18D; &#xD53C;&#xB4DC;&#xBC31; &#xB8E8;&#xD504;&#xB294; &#xC11C;&#xB85C; &#xB2E4;&#xB978; &#xC218;&#xC900;&#xC5D0;&#xC11C; cyclin B Cdk1 &#xD65C;&#xC131;&#xD654;&#xB97C; &#xAC15;&#xD654; &#xC874;&#xC7AC; &#xC124;&#xBA85; &#xD588;&#xB2E4;. &#xC774; &#xAC80;&#xD1A0;&#xC5D0;&#xC11C; &#xC6B0;&#xB9AC;&#xB294; &#xB2E4;&#xB978; &#xC870;&#xAC74; &#xD558;&#xC5D0;&#xC11C; &#xADF8;&#xB4E4;&#xC758; &#xD65C;&#xC131;&#xD654; &#xC2DC;&#xAE30;&#xC640; &#xC778;&#xAC04; &#xC138;&#xD3EC; mitotic &#xD56D;&#xBAA9;&#xC744; &#xC81C;&#xC5B4; &#xD558;&#xAE30; &#xC5B4;&#xB824;&#xC6B4; &#xD2B8;&#xB9AC;&#xAC70;&#xC758; &#xAC00;&#xB2A5;&#xD55C; id cyclin B Cdk1 &#xD65C;&#xC131;&#xD654;&#xB97C; &#xC870;&#xC808; &#xD558;&#xB294; &#xB2E4;&#xC591; &#xD55C; &#xD53C;&#xB4DC;&#xBC31; &#xB8E8;&#xD504;&#xC758; &#xC5ED;&#xD560; &#xB17C;&#xC758;.

&#xC720;&#xC0AC; &#xBD84;&#xC5F4;&#xC744; &#xC785;&#xB825; &#xACB0;&#xC815;: &#xD53C;&#xB4DC;&#xBC31; &#xBC0F; mitotic &#xD56D;&#xBAA9; &#xB124;&#xD2B8;&#xC6CC;&#xD06C;&#xC5D0; &#xC911;&#xBCF5;.

La decisi&#xF3;n de entrar en la mitosis est&#xE1; mediada por una red de prote&#xED;nas que regulan la activaci&#xF3;n de la ciclina B-Cdk1 complejo. Dentro de esta red, varios bucles de retroalimentaci&#xF3;n positiva puede amplificar la ciclina B-Cdk1 de activaci&#xF3;n para asegurar el compromiso completo a un estado mit&#xF3;tico una vez que la decisi&#xF3;n de entrar en la mitosis se ha hecho. Sin embargo, se est&#xE1; acumulando evidencia de que varios componentes de los circuitos de retroalimentaci&#xF3;n son redundantes para la ciclina B-Cdk1 activaci&#xF3;n durante la divisi&#xF3;n celular normal. Sin embargo, se definen los circuitos de retroalimentaci&#xF3;n convertido en algo esencial para promover la entrada mit&#xF3;tico, cuando la progresi&#xF3;n del ciclo celular normal es perturbado. Los datos recientes han demostrado que al menos tres Plk1 que dependen de los bucles de retroalimentaci&#xF3;n existe de que mejorar la ciclina B-Cdk1 de activaci&#xF3;n en diferentes niveles. En esta revisi&#xF3;n, se discute el papel de los diferentes circuitos de retroalimentaci&#xF3;n que regulan la ciclina B-Cdk1 la activaci&#xF3;n en diferentes condiciones, el momento de su activaci&#xF3;n, y la posible identidad de el gatillo dif&#xED;cil que controla la entrada mit&#xF3;tico en las c&#xE9;lulas humanas.

La decisi&#xF3;n de entrar en mitosis: Comentarios y redundancia en la red de entrada mit&#xF3;tico

&#x52A8;&#x7269;&#x7EC6;&#x80DE;&#x542F;&#x52A8;&#x540E;&#x671F;&#x53D1;&#x75C5;&#xFF0C;&#x4E0E;&#x5E76;&#x884C;&#x7684;&#x80DE;&#x8D28;&#x5206;&#x88C2;&#x65F6;&#x8089;&#x73AF;&#x88C5;&#x914D;&#x548C;&#x672C;&#x5730;&#x5316;&#x6D3B;&#x5316;&#x7684;&#x5C0F;&#x86CB;&#x767D; RhoA&#xFF0C;&#x5F15;&#x8D77;&#x5288;&#x88C2;&#x6C9F;&#x901A;&#x8FC7;&#x4E0D;&#x7545;&#x3002;&#x6C9F;&#x5F62;&#x6210;&#x4F9D;&#x8D56;&#x4E8E;&#x4F4D;&#x7F6E;&#x63D0;&#x793A;&#x63D0;&#x4F9B;&#x7684;&#x540E;&#x671F;&#x4E3B;&#x8F74;&#x5FAE;&#x7BA1; (MTs)&#xFF0C;&#x4F46;&#x8FD9;&#x79CD;&#x7EBF;&#x7D22;&#x5982;&#x4F55;&#x751F;&#x6210;&#x4ECD;&#x4E0D;&#x6E05;&#x695A;&#x3002;&#x6211;&#x4EEC;&#x4F7F;&#x7528;&#x5316;&#x5B66;&#x9057;&#x4F20;&#x5B66;&#x6765;&#x5B9E;&#x73B0;&#x65F6;&#x95F4;&#x548C;&#x7A7A;&#x95F4;&#x7684;&#x63A7;&#x5236;&#xFF0C;&#x663E;&#x793A;&#x7684; Polo &#x6837;&#x6FC0;&#x9176; 1 (Plk1) &#x6210;&#x548C;&#x5176;&#x5730;&#x65B9;&#x7684;&#x78F7;&#x9178;&#x5316;&#x5C71;&#x7ED1;&#x5B9A;&#x886C;&#x5E95;&#x7684;&#x81EA;&#x7EC4;&#x7EC7;&#x7684;&#x4EA4;&#x4ED8;&#x6B64;&#x6C9F;&#x8BF1;&#x5BFC;&#x4FE1;&#x53F7;&#x4EA7;&#x751F;&#x7684;&#x5173;&#x952E;&#x3002;Plk1 &#x65F6;&#x6D3B;&#x52A8;&#xFF0C;&#x4F46;&#x4E0D;&#x80FD;&#x628A;&#x76EE;&#x6807;&#x672C;&#x8EAB;&#x8FD9;&#x4E2A;&#x8D64;&#x9053;&#x7684;&#x91CC;&#x7A0B;&#x7891;&#xFF0C;&#x8FD9;&#x4E24;&#x79CD;&#x76AE;&#x8D28; RhoA &#x5F81;&#x8058;&#x548C;&#x6C9F;&#x611F;&#x5E94;&#x672A;&#x80FD;&#x51FA;&#x73B0;&#xFF0C;&#x56E0;&#x6B64;&#x56DE;&#x987E;&#x540E;&#x671F;&#x7279;&#x5B9A; Plk1 &#x6291;&#x5236;&#x4F5C;&#x7528;&#x7684;&#x5F71;&#x54CD;&#x3002;&#x6211;&#x4EEC;&#x4F7F;&#x7528;&#x4E32;&#x8054;&#x8D28;&#x8C31;&#x548C; phosphospecific &#x6297;&#x4F53;&#xFF0C;&#x53D1;&#x73B0; Plk1 &#x7ED1;&#x5B9A;&#xFF0C;&#x5E76;&#x4E14;&#x76F4;&#x63A5; phosphorylates centralspindlin &#xFF08;&#x4E5F;&#x79F0;&#x4E3A; MgcRacGAP&#xFF09; &#x5728;&#x6210;&#x7B2C; HsCYK-4 &#x4E9A;&#x57FA;&#x3002;&#x5728;&#x4E1D;&#x6C28;&#x9178; 157&#xFF0C;&#x6B64;&#x4FEE;&#x6539;&#x521B;&#x5EFA; BRCT &#x91CD;&#x590D;&#x7684; Rho GTP &#x4EA4;&#x6D41;&#x56E0;&#x5B50; Ect2 &#x7684;&#x4E32;&#x8054;&#x4E3B;&#x8981;&#x505C;&#x9760;&#x7AD9;&#x70B9;&#x3002;&#x8868;&#x793A;&#x53EA;&#x6709; nonphosphorylatable HsCYK-4 &#x7684;&#x7A97;&#x4F53;&#x7684;&#x7EC6;&#x80DE;&#x672A;&#x80FD;&#x5728;&#x6210;&#x672C;&#x5730;&#x5316; Ect2 &#x548C;&#x5288;&#x88C2;&#x6C9F;&#x5F62;&#x6210;&#xFF0C;&#x8FD9;&#x610F;&#x5473;&#x7740; HsCYK 4 Plk1 &#x7684;&#x901F;&#x7387;&#x9650;&#x5236;&#x4E0A;&#x6E38;&#x7684;&#x76EE;&#x6807; RhoA &#x5728;&#x4E25;&#x91CD;&#x524A;&#x5F31;&#x4E86;&#x3002;&#x76F8;&#x53CD;&#xFF0C;&#x4E2D;&#x7EE7;&#x5230; HsCYK 4 &#x5141;&#x8BB8;&#x5411;&#x7A97;&#x4F53;&#x5C3D;&#x7BA1;&#x5168;&#x7403;&#x7684;&#x5355;&#x5143;&#x683C;&#x4E2D;&#x7684;&#x6240;&#x6709;&#x5176;&#x4ED6; Plk1 &#x5206;&#x5B50;&#x6291;&#x5236;&#x76B1;&#x7EB9; Plk1 &#x6291;&#x5236;&#x5242;&#x8010;&#x836F;&#x57FA;&#x56E0;&#x3002;&#x6211;&#x4EEC;&#x7684;&#x7814;&#x7A76;&#x7ED3;&#x679C;&#x7167;&#x4EAE;&#x4EBA;&#x7C7B;&#x7EC6;&#x80DE;&#x80DE;&#x8D28;&#x5206;&#x88C2;&#x7684;&#x5F00;&#x59CB;&#x7684;&#x4E24;&#x4E2A;&#x5173;&#x952E;&#x673A;&#x5236;&#xFF0C;&#x5E76;&#x8BF4;&#x660E;&#x5316;&#x5B66;&#x9057;&#x4F20;&#x5B66;&#x63A2;&#x9488;&#x5728;&#x65F6;&#x95F4;&#x548C;&#x7A7A;&#x95F4;&#x8FD9;&#x79CD;&#x76D1;&#x7BA1;&#x7684;&#x6743;&#x529B;&#x3002;

Plk1 &#x81EA;&#x7EC4;&#x7EC7;&#x548C;&#x9884;&#x5145;&#x78F7;&#x9178;&#x5316;&#x7684; HsCYK 4 &#x4E3B;&#x8F74;&#x6210;&#x5728;&#x89C4;&#x7BA1;&#x53F8;&#x5728;&#x4EBA;&#x7C7B;&#x7EC6;&#x80DE;&#x4E2D;&#x7684;&#x53D1;&#x75C5;&#x3002;

Animal cells initiate cytokinesis in parallel with anaphase onset, when an actomyosin ring assembles and constricts through localized activation of the small GTPase RhoA, giving rise to a cleavage furrow. Furrow formation relies on positional cues provided by anaphase spindle microtubules (MTs), but how such cues are generated remains unclear. Using chemical genetics to achieve both temporal and spatial control, we show that the self-organized delivery of Polo-like kinase 1 (Plk1) to the midzone and its local phosphorylation of a MT-bound substrate are critical for generating this furrow-inducing signal. When Plk1 was active but unable to target itself to this equatorial landmark, both cortical RhoA recruitment and furrow induction failed to occur, thus recapitulating the effects of anaphase-specific Plk1 inhibition. Using tandem mass spectrometry and phosphospecific antibodies, we found that Plk1 binds and directly phosphorylates the HsCYK-4 subunit of centralspindlin (also known as MgcRacGAP) at the midzone. At serine 157, this modification creates a major docking site for the tandem BRCT repeats of the Rho GTP exchange factor Ect2. Cells expressing only a nonphosphorylatable form of HsCYK-4 failed to localize Ect2 at the midzone and were severely impaired in cleavage furrow formation, implying that HsCYK-4 is Plk1's rate-limiting target upstream of RhoA. Conversely, tethering an inhibitor-resistant allele of Plk1 to HsCYK-4 allowed furrows to form despite global inhibition of all other Plk1 molecules in the cell. Our findings illuminate two key mechanisms governing the initiation of cytokinesis in human cells and illustrate the power of chemical genetics to probe such regulation both in time and space.

Plk1 self-organization and priming phosphorylation of HsCYK-4 at the spindle midzone regulate the onset of division in human cells.

Les cellules animales lancer cytocin&egrave;se parall&egrave;lement &agrave; l&#39;apparition de l&#39;anaphase, lorsqu&#39;une bague d&#39;actomyosine assemble et resserre &agrave; travers l&#39;activation locale de la petite GTPase RhoA, donnant lieu &agrave; un sillon de clivage. Sillon formation repose sur positionnels indices fournis par les microtubules du fuseau de l&#39;anaphase (MTs), mais comment ces signaux sont g&eacute;n&eacute;r&eacute;s reste incertaine. En utilisant la g&eacute;n&eacute;tique chimique pour obtenir un contr&ocirc;le spatial et temporel, nous montrons que la livraison auto-organis&eacute;s de Polo-like kinase 1 (Plk1) pour le milieu et sa phosphorylation locale d&#39;un substrat li&eacute; aux MT sont essentiels pour g&eacute;n&eacute;rer ce signal induisant la raie de semis. Quand Plk1 &eacute;tait actif mais impossible de cibler elle-m&ecirc;me &agrave; ce point de rep&egrave;re &eacute;quatorial, deux corticales RhoA recrutement et sillon l&#39;induction n&#39;a pas se produire, reprenant ainsi les effets de l&#39;inhibition de Plk1 sp&eacute;cifiques &agrave; l&#39;anaphase. &Agrave; l&#39;aide de spectrom&eacute;trie de masse et les anticorps phosphospecific, nous avons constat&eacute; que Plk1 se lie et phosphoryle directement la sous-unit&eacute; 4-HsCYK de centralspindlin (&eacute;galement connu sous le nom de MgcRacGAP) dans le milieu. &Agrave; s&eacute;rine 157, cette modification cr&eacute;e un site majeur d&#39;amarrage pour le tandem que BRCT r&eacute;p&egrave;te du facteur Rho GTP change Ect2. Les cellules n&#39;exprimant qu&#39;une forme de nonphosphorylatable de HsCYK-4 n&#39;a pas pu localiser le Ect2 dans le milieu et ont gravement port&eacute; atteinte dans la formation de sillon de clivage, ce qui implique que HsCYK-4 est limitante cible de Plk1 en amont de RhoA. &Agrave; l&#39;inverse, attacher un all&egrave;le r&eacute;sistants aux inhibiteur de Plk1 &agrave; HsCYK-4 a permis des sillons de forme malgr&eacute; une inhibition globale de toutes les autres mol&eacute;cules de Plk1 dans la cellule. Nos r&eacute;sultats s&#39;allume deux m&eacute;canismes essentiels r&eacute;gissant l&#39;ouverture de la cytokin&egrave;se dans les cellules humaines et illustrent la puissance de la g&eacute;n&eacute;tique chimique pour sonder cette r&eacute;glementation aussi bien dans le temps et l&#39;espace.

Auto-organisation de Plk1 et phosphorylation d&#39;amor&ccedil;age du HsCYK-4 au milieu de l&#39;axe r&eacute;glementent l&#39;apparition de la division dans les cellules humaines.

Tierzellen initiieren Zellteilung parallel zu Beginn der Anaphase, wenn ein Actomyosin Ring montiert und durch lokalisierte Aktivierung von die kleine GTPase RhoA, was zu einer Trennungsfurche verengt. Furche Bildung st&uuml;tzt sich auf positionelle Hinweise bereitgestellt durch Anaphase Spindel Mikrotubuli (MTs), aber es bleibt unklar, wie solche Signale generiert werden. Verwenden chemische Genetik, um sowohl zeitliche und r&auml;umliche Kontrolle zu erreichen, zeigen wir, dass die selbstorganisierte Lieferung von Polo-Like Kinase 1 (Plk1) die Midzone und seine lokalen Phosphorylierung MT gebundene Substrat sind entscheidend f&uuml;r die generiert diese Furche-induzierende Signals. Als Plk1 aktiv, aber nicht in der Lage, selbst diese &auml;quatorialen Wahrzeichen Ziel war, konnte beide kortikale RhoA Rekrutierung und Furche Induktion auftreten, damit die Auswirkungen der Anaphase-spezifische Plk1 Hemmung Biodiversit&auml;tsmonitoring. Mit Tandem-Massenspektrometrie und Phosphospecific Antik&ouml;rper, fanden wir, dass Plk1 bindet und direkt die HsCYK-4-Untereinheit des Centralspindlin (auch bekannt als MgcRacGAP) an die Midzone phosphorylates. Am Serin 157 erstellt diese &Auml;nderung eine gro&szlig;e docking-Website f&uuml;r das Tandem, wie, das BRCT des Faktors Austausch Rho GTP Ect2 wiederholt wird. Zellen Ausdr&uuml;cken nur eine Nonphosphorylatable Form des HsCYK-4 Fehler beim Ect2 an den Midzone zu lokalisieren und wurden in Spaltung Furche-Formation, was bedeutet, dass HsCYK-4 Plk1s geschwindigkeitsbestimmende Ziel stromaufw&auml;rts von RhoA stark beeintr&auml;chtigt. Umgekehrt Anbindehaltung ein Inhibitor-resistenten Allel des Plk1 bis HsCYK-4 erlaubt Furchen zu bilden trotz globaler Hemmung anderer Plk1-Molek&uuml;le in der Zelle. Unsere Ergebnisse beleuchten zwei Schl&uuml;sselmechanismen f&uuml;r die Einleitung der Zellteilung in den menschlichen Zellen und illustrieren die Macht der chemische Genetik zu dieser Verordnung, die sowohl in Zeit und Raum zu sondieren.

Plk1 Selbstorganisation und Grundierung Phosphorylierung von HsCYK-4 an die Spindel Midzone regulieren den Ausbruch der Division in den menschlichen Zellen.

Cellule animali avviano Citocinesi in parallelo con l&#39;esordio di anafase, quando un anello di actomiosina assembla e restringe attraverso l&#39;attivazione localizzata del piccolo GTPase RhoA, dando luogo a un solco di clivaggio. Formazione del solco si basa su posizionali spunti forniti da microtubuli di mandrino anafase (MTs), ma come tali spunti sono generati rimane poco chiaro. Utilizzando chimica genetica per ottenere un controllo temporale e spaziale, mostriamo che la consegna auto-organizzata del Polo-like chinasi 1 (Plk1) per la midzone e la sua fosforilazione locale di un substrato associato a MT sono critici per la generazione di questo segnale che inducono solco. Quando Plk1 era attivo ma incapace di target stesso a questa pietra miliare equatoriale, entrambi induzione reclutamento e solco corticale RhoA impossibile a verificarsi, quindi ricapitolando gli effetti di inibizione Plk1 anafase-specifici. Usando gli anticorpi phosphospecific e spettrometria di massa tandem, abbiamo scoperto che Plk1 si lega e fosforila direttamente la subunit&agrave; HsCYK-4 di centralspindlin (noto anche come MgcRacGAP) presso il midzone. A serina 157, questa modifica consente di creare un importante sito di aggancio per il tandem che BRCT ripete il fattore di scambio GTP Rho Ect2. Le cellule che esprimono solo una forma di nonphosphorylatable di HsCYK-4 non &egrave; riuscito a localizzare Ect2 presso il midzone e sono stati gravemente compromessa in formazione di solco di clivaggio, implicando che HsCYK-4 &egrave; rate-limiting destinazione di Plk1 a Monte di RhoA. Al contrario, legare un allele inibitore-resistente di Plk1 di HsCYK-4 permesso solchi a forma nonostante l&#39;inibizione globale di tutte le altre molecole Plk1 nella cella. I nostri risultati illuminano due meccanismi fondamentali che regolano l&#39;avvio della Citocinesi in cellule umane e illustrano la potenza della chimica genetica per sondare tale regolamento sia nel tempo e nello spazio.

Plk1 auto-organizzazione e fosforilazione di innesco di HsCYK-4 presso il midzone mandrino regolano l&#39;insorgenza della divisione nelle cellule umane.

&#x30A2;&#x30AF;&#x30C8;&#x30DF;&#x30AA;&#x30B7;&#x30F3;&#x306E;&#x30EA;&#x30F3;&#x30B0;&#x3092;&#x30A2;&#x30BB;&#x30F3;&#x30D6;&#x30EB;&#x3057;&#x3001;&#x6E9D;&#x90E8;&#x306B;&#x4E0A;&#x6607;&#x3092;&#x4E0E;&#x3048;&#x3066;&#x3001;&#x5C0F;&#x3055;&#x306A;&#x306E; gtp &#x30A2;&#x30FC;&#x30BC; RhoA &#x306E;&#x30ED;&#x30FC;&#x30AB;&#x30E9;&#x30A4;&#x30BA;&#x3055;&#x308C;&#x305F;&#x6D3B;&#x6027;&#x5316;&#x3092;&#x4ECB;&#x3057;&#x3066;&#x53CE;&#x7E2E;&#x3059;&#x308B;&#x3068;&#x304D;&#x52D5;&#x7269;&#x7D30;&#x80DE;&#x5206;&#x88C2;&#x3068;&#x4E26;&#x884C;&#x3057;&#x3066;&#x5F8C;&#x671F;&#x767A;&#x75C7;&#x3092;&#x958B;&#x59CB;&#x3057;&#x307E;&#x3059;&#x3002;&#x6E9D;&#x5F62;&#x6210;&#x5F8C;&#x671F;&#x30B9;&#x30D4;&#x30F3;&#x30C9;&#x30EB;&#x5FAE;&#x5C0F;&#x7BA1; (MTs) &#x3092;&#x63D0;&#x4F9B;&#x4F4D;&#x7F6E;&#x624B;&#x304C;&#x304B;&#x308A;&#x304C;&#x4F9D;&#x5B58;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x304C;&#x3001;&#x3053;&#x306E;&#x3088;&#x3046;&#x306A;&#x624B;&#x304C;&#x304B;&#x308A;&#x306E;&#x751F;&#x6210;&#x65B9;&#x6CD5;&#x306F;&#x4E0D;&#x660E;&#x306E;&#x307E;&#x307E;&#x3002;&#x5316;&#x5B66;&#x907A;&#x4F1D;&#x5B66;&#x306F;&#x3001;&#x6642;&#x9593;&#x3068;&#x7A7A;&#x9593;&#x306E;&#x5236;&#x5FA1;&#x3092;&#x5B9F;&#x73FE;&#x3059;&#x308B;&#x3092;&#x4F7F;&#x7528;&#x3057;&#x3066;&#x3001;&#x81EA;&#x5DF1;&#x7D44;&#x7E54;&#x5316;&#x306E;&#x914D;&#x4FE1;&#x306E;&#x30DD;&#x30ED;&#x69D8;&#x30AD;&#x30CA;&#x30FC;&#x30BC; 1 &#xFF08;Plk1&#xFF09;&#x3001;midzone &#x3068;&#x305D;&#x306E;&#x30ED;&#x30FC;&#x30AB;&#x30EB; &#x30EA;&#x30F3;&#x9178; MT &#x30D0;&#x30A4;&#x30F3;&#x30C9;&#x57FA;&#x677F;&#x304C;&#x3053;&#x306E;&#x7566;&#x9593;&#x8A98;&#x5C0E;&#x4FE1;&#x53F7;&#x3092;&#x751F;&#x6210;&#x3059;&#x308B;&#x305F;&#x3081;&#x306B;&#x91CD;&#x8981;&#x3067;&#x3042;&#x308B;&#x3053;&#x3068;&#x3092;&#x793A;&#x3057;&#x307E;&#x3059;&#x3002;Plk1 &#x304C;&#x30A2;&#x30AF;&#x30C6;&#x30A3;&#x30D6;&#x3060;&#x304C;&#x3067;&#x304D;&#x307E;&#x305B;&#x3093;&#x81EA;&#x4F53;&#x3053;&#x306E;&#x8D64;&#x9053;&#x306E;&#x30E9;&#x30F3;&#x30C9;&#x30DE;&#x30FC;&#x30AF;&#x3092;&#x5BFE;&#x8C61;&#x3068;&#x3059;&#x308B;&#x9803;&#x3001;&#x4E21;&#x65B9;&#x306E;&#x76AE;&#x8CEA; RhoA &#x8A98;&#x5C0E;&#x52DF;&#x96C6;&#x304A;&#x3088;&#x3073;&#x6E9D;&#x304C;&#x767A;&#x751F;&#x3059;&#x308B;&#x306B;&#x306F;&#x3001;&#x3053;&#x3046;&#x3057;&#x3066;&#x7279;&#x5B9A;&#x306E;&#x5F8C;&#x671F; Plk1 &#x963B;&#x5BB3;&#x306E;&#x5F71;&#x97FF;&#x305A;&#x3064;&#x3092;&#x6458;&#x8A18;&#x3067;&#x3057;&#x305F;&#x3002;&#x30BF;&#x30F3;&#x30C7;&#x30E0;&#x8CEA;&#x91CF;&#x5206;&#x6790;&#x6CD5;&#x3068; phosphospecific &#x306E;&#x6297;&#x4F53;&#x3092;&#x4F7F;&#x7528;&#x3057;&#x3066;&#x3001;Plk1 &#x304C;&#x7D50;&#x5408;&#x3057;&#x3001;&#x76F4;&#x63A5; centralspindlin &#xFF08;&#x5225;&#x540D; MgcRacGAP&#xFF09;&#x3001;midzone &#x3067;&#x306E; HsCYK 4 &#x30B5;&#x30D6;&#x30E6;&#x30CB;&#x30C3;&#x30C8;&#x3067;&#x3042;&#x308B;&#x3053;&#x3068;&#x3092;&#x767A;&#x898B;&#x3057;&#x307E;&#x3057;&#x305F;&#x3002;&#x30BB;&#x30EA;&#x30F3; 157 &#x3067;&#x3053;&#x306E;&#x5909;&#x66F4; BRCT Rho GTP &#x4EA4;&#x63DB;&#x56E0;&#x5B50; Ect2 &#x306E;&#x7E70;&#x308A;&#x8FD4;&#x3055;&#x308C;&#x308B;&#x30BF;&#x30F3;&#x30C7;&#x30E0;&#x306E;&#x305F;&#x3081;&#x306E;&#x4E3B;&#x8981;&#x306A;&#x30C9;&#x30C3;&#x30AD;&#x30F3;&#x30B0; &#x30B5;&#x30A4;&#x30C8;&#x3092;&#x4F5C;&#x6210;&#x3057;&#x307E;&#x3059;&#x3002;HsCYK 4 &#x306E;&#x5F62;&#x3067;&#x306E;&#x307F; nonphosphorylatable &#x3092;&#x767A;&#x73FE;&#x3059;&#x308B;&#x7D30;&#x80DE;&#x306F; Ect2 midzone &#x3067;&#x30ED;&#x30FC;&#x30AB;&#x30E9;&#x30A4;&#x30BA;&#x3067;&#x304D;&#x307E;&#x305B;&#x3093;&#x3067;&#x3057;&#x305F;&#x3057;&#x3001;HsCYK 4 Plk1 &#x306E;&#x30EC;&#x30FC;&#x30C8;&#x5236;&#x9650;&#x4E0A;&#x6D41;&#x306E;&#x30BF;&#x30FC;&#x30B2;&#x30C3;&#x30C8; RhoA &#x3067;&#x3059;&#x793A;&#x5506;&#x958B;&#x88C2;&#x6E9D;&#x5F62;&#x6210;&#x3067;&#x306F;&#x3001;&#x3072;&#x3069;&#x304F;&#x640D;&#x306A;&#x308F;&#x308C;&#x307E;&#x3057;&#x305F;&#x3002;&#x9006;&#x306B;&#x3001;&#x3059;&#x308B;&#x963B;&#x5BB3;&#x5264;&#x8010;&#x6027;&#x306E;&#x5BFE;&#x7ACB;&#x907A;&#x4F1D;&#x5B50;&#x306E; Plk1 HsCYK 4 &#x5C0F;&#x6E9D;&#x4ED6;&#x306E;&#x3059;&#x3079;&#x3066;&#x306E; Plk1 &#x5206;&#x5B50;&#x7D30;&#x80DE;&#x5185;&#x306E;&#x30B0;&#x30ED;&#x30FC;&#x30D0;&#x30EB;&#x6291;&#x5236;&#x306B;&#x3082;&#x304B;&#x304B;&#x308F;&#x3089;&#x305A;&#x30D5;&#x30A9;&#x30FC;&#x30E0;&#x306B;&#x8A31;&#x53EF;&#x3059;&#x308B;&#x30C6;&#x30B6;&#x30EA;&#x30F3;&#x30B0;&#x3002;&#x6211;&#x3005; &#x306E;&#x8ABF;&#x67FB;&#x7D50;&#x679C;&#x306F;&#x3072;&#x3068;&#x7D30;&#x80DE;&#x306E;&#x7D30;&#x80DE;&#x8CEA;&#x5206;&#x88C2;&#x306E;&#x958B;&#x59CB;&#x3092;&#x652F;&#x914D;&#x3059;&#x308B; 2 &#x3064;&#x306E;&#x4E3B;&#x8981;&#x30E1;&#x30AB;&#x30CB;&#x30BA;&#x30E0;&#x3092;&#x7167;&#x3089;&#x3059;&#x3057;&#x3001;&#x6642;&#x9593;&#x3068;&#x7A7A;&#x9593;&#x306E;&#x4E21;&#x65B9;&#x3067;&#x305D;&#x306E;&#x3088;&#x3046;&#x306A;&#x898F;&#x5236;&#x3092;&#x30D7;&#x30ED;&#x30FC;&#x30D6;&#x3059;&#x308B;&#x5316;&#x5B66;&#x907A;&#x4F1D;&#x5B66;&#x306E;&#x529B;&#x3092;&#x793A;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;

Plk1 &#x81EA;&#x5DF1;&#x7D44;&#x7E54;&#x5316;&#x3068;&#x30B9;&#x30D4;&#x30F3;&#x30C9;&#x30EB; midzone &#x3067; HsCYK 4 &#x306E;&#x30D7;&#x30E9;&#x30A4;&#x30DF;&#x30F3;&#x30B0;&#x306E;&#x30EA;&#x30F3;&#x9178;&#x5316;&#x306F;&#x3001;&#x4EBA;&#x9593;&#x306E;&#x7D30;&#x80DE;&#x5206;&#x88C2;&#x306E;&#x767A;&#x75C7;&#x3092;&#x8ABF;&#x7BC0;&#x3057;&#x307E;&#x3059;&#x3002;

&#xB3D9;&#xBB3C; &#xC138;&#xD3EC; actomyosin &#xB9C1; &#xC870;&#xB9BD; &#xD558; &#xACE0; &#xC720; &#xBC29; &#xC0AC;&#xC774;&#xC758; &#xACE8;&#xC9DC;&#xAE30; &#xBA85;&#xC529;&#xC744; &#xC57C; &#xAE30;&#xD55C; &#xC791;&#xC740; GTPase Rhoa&#xC758; &#xC9C0;&#xC5ED;&#xD654; &#xB41C; &#xD65C;&#xC131;&#xD654;&#xB97C; &#xD1B5;&#xD574; &#xC218;&#xCD95; cytokinesis anaphase &#xAC1C;&#xC2DC;&#xC640; &#xB3D9;&#xC2DC;&#xC5D0; &#xC2DC;&#xC791; &#xD569;&#xB2C8;&#xB2E4;. &#xBA85;&#xC529; &#xD615;&#xC131; anaphase &#xC2A4;&#xD540; &#xB4E4; microtubules (MTs)&#xC5D0;&#xC11C; &#xC81C;&#xACF5; &#xD558;&#xB294; &#xC704;&#xCE58; &#xC2E0;&#xD638;&#xC5D0; &#xC758;&#xC874; &#xD558;&#xC9C0;&#xB9CC; &#xBD88;&#xBD84;&#xBA85; &#xB0A8;&#xC544; &#xAC19;&#xC740; &#xC2E0;&#xD638; &#xC0DD;&#xC131; &#xBC29;&#xBC95;. &#xD654;&#xD559; &#xC720;&#xC804;&#xD559;&#xC744; &#xC0AC;&#xC6A9; &#xD558; &#xC5EC; &#xC2DC;&#xAC04;&#xC801; &#xBC0F; &#xACF5;&#xAC04;&#xC801; &#xC81C;&#xC5B4;&#xB97C; &#xB2EC;&#xC131; &#xD558;&#xAE30; &#xC704;&#xD574;, &#xC6B0;&#xB9AC;&#xB294; &#xD3F4;&#xB85C; &#xAC19;&#xC740; &#xD0A4; 1 (Plk1)&#xB294; midzone&#xC640; &#xB85C;&#xCEEC;&#xC758; &#xC778; &#xC0B0;&#xD654;&#xB97C; &#xB9C8;&#xC6B4;&#xD2B8; &#xBC14;&#xC778;&#xB529;&#xB41C; &#xAE30;&#xD310;&#xC758; &#xC790;&#xAE30; &#xC804;&#xB2EC;&#xC774; &#xBA85;&#xC529; &#xC720;&#xB3C4; &#xC2E0;&#xD638;&#xB97C; &#xC0DD;&#xC131; &#xD558;&#xAE30; &#xC704;&#xD55C; &#xC911;&#xC694; &#xD55C;&#xB294; &#xBCF4;&#xC5EC;&#xC90D;&#xB2C8;&#xB2E4;. Plk1 &#xD65C;&#xC131; &#xC0C1;&#xD0DC; &#xC774;&#xC9C0;&#xB9CC;&#xC774; &#xC801;&#xB3C4; &#xB79C;&#xB4DC;&#xB9C8;&#xD06C;&#xB97C; &#xB300;&#xC0C1;&#xC73C;&#xB85C; &#xC790;&#xCCB4; &#xC218; &#xC5C6;&#xC744; &#xB54C;, &#xB450; &#xD53C; &#xC9C8; RhoA &#xBAA8;&#xC9D1; &#xBC0F; &#xBA85;&#xC529; &#xC720;&#xB3C4; &#xB530;&#xB77C;&#xC11C; &#xC5C5;&#xACFC; anaphase &#xD2B9;&#xC815; Plk1 &#xC800;&#xD574; &#xD6A8;&#xACFC; &#xBC1C;&#xC0DD;&#xC744; &#xBABB;&#xD588;&#xC2B5;&#xB2C8;&#xB2E4;. &#xD0E0;&#xB364; &#xC9C8;&#xB7C9; &#xBD84;&#xC11D; &#xBC0F; phosphospecific &#xD56D; &#xCCB4;&#xB97C; &#xC0AC;&#xC6A9; &#xD558; &#xC5EC; plk1&#xC5D0; &#xBC14;&#xC778;&#xB529;&#xD569;&#xB2C8;&#xB2E4; &#xBC0F; &#xC9C1;&#xC811; phosphorylates centralspindlin (MgcRacGAP &#xB77C;&#xACE0;&#xB3C4; &#xD568;)&#xB97C; midzone&#xC758; HsCYK-4 &#xC18C; &#xB2E8;&#xC704; &#xCC3E;&#xC558;&#xC2B5;&#xB2C8;&#xB2E4;. &#xB5A0;&#xB4E4;&#xACE0; 157&#xC5D0;&#xC774; &#xC218;&#xC815; BRCT &#xBC18;&#xBCF5; Rho GTP exchange &#xC694;&#xC18C; ect2&#xC758; &#xD611;&#xB3D9;&#xC5D0; &#xB300; &#xD55C; &#xC8FC;&#xC694; &#xB3C4;&#xD0B9; &#xC0AC;&#xC774;&#xD2B8;&#xB97C; &#xB9CC;&#xB4ED;&#xB2C8;&#xB2E4;. HsCYK 4 nonphosphorylatable &#xC591;&#xC2DD;&#xB9CC;&#xC744; &#xD45C;&#xD604; &#xD558;&#xB294; &#xC140;&#xC740; midzone&#xC5D0;&#xC11C; ect2&#xB97C; &#xC9C0;&#xC5ED;&#xD654; &#xD558;&#xB294; &#xB370; &#xC2E4;&#xD328; &#xD558; &#xACE0; HsCYK 4 plk1&#xC758; &#xC18D;&#xB3C4; &#xC81C;&#xD55C; &#xB300;&#xC0C1; &#xC0C1;&#xB958; Rhoa&#xC758; &#xC554;&#xC2DC; &#xBD84;&#xC5F4; &#xBA85;&#xC529; &#xD615;&#xC131;&#xC5D0; &#xC7A5;&#xC560;&#xAC00; &#xC2EC;&#xAC01; &#xD588;&#xB2E4;. &#xBC18;&#xB300;&#xB85C;, Plk1 HsCYK-4 &#xC140;&#xC5D0; &#xB2E4;&#xB978; &#xBAA8;&#xB4E0; Plk1 &#xBD84;&#xC790;&#xC758; &#xAE00;&#xB85C;&#xBC8C; &#xC5B5;&#xC81C;&#xC5D0;&#xB3C4; &#xBD88;&#xAD6C; &#xD558; &#xACE0; &#xD615;&#xD0DC;&#xB97C; &#xACE0; &#xB791;&#xC744; &#xD5C8;&#xC6A9;&#xC758; &#xC5B5;&#xC81C;&#xC81C; &#xBC29;&#xC9C0; &#xB300;&#xB9BD; &#xC720;&#xC804;&#xC790;&#xB97C; &#xBC27;&#xC904;&#xB85C; &#xB9E4;&#xAE30;. &#xC6B0;&#xB9AC;&#xC758; &#xC5F0;&#xAD6C; &#xACB0;&#xACFC; &#xC778;&#xAC04; &#xC138;&#xD3EC;&#xC5D0;&#xC11C; cytokinesis&#xC758; &#xAC1C;&#xC2DC; &#xD1B5;&#xCE58; &#xB450; &#xAC00;&#xC9C0; &#xC8FC;&#xC694; &#xBA54;&#xCEE4;&#xB2C8;&#xC998;&#xC744; &#xBC1D;&#xD788;&#xB294; &#xACE0; &#xD654;&#xD559; &#xC720;&#xC804;&#xD559;&#xC758; &#xD798;&#xC744; &#xBAA8;&#xB450; &#xC2DC;&#xAC04;&#xACFC; &#xACF5;&#xAC04;&#xC5D0; &#xC774;&#xB7EC;&#xD55C; &#xADDC;&#xC815;&#xC744; &#xD504;&#xB85C;&#xBE0C;&#xB97C; &#xBCF4;&#xC5EC;&#xC90D;&#xB2C8;&#xB2E4;.

Plk1 &#xC790;&#xAE30;&#xC640; &#xC2A4;&#xD540; &#xB4E4; midzone&#xC5D0;&#xC11C; HsCYK-4&#xC758; &#xC560; &#xBC8C; &#xCE60; &#xC778; &#xC0B0;&#xD654; &#xC778;&#xAC04;&#xC758; &#xC138;&#xD3EC; &#xBD84;&#xC5F4;&#xC758; &#xBC1C;&#xBCD1;&#xC744; &#xC870;&#xC808;.

Las c&eacute;lulas animales inician citocinesis en paralelo con el inicio de la anafase, cuando un anillo de actomyosin monta y esto constri&ntilde;e a trav&eacute;s de la activaci&oacute;n localizada de la peque&ntilde;a GTPase RhoA, dando lugar a un surco de escote. Formaci&oacute;n del surco se basa en posicional, proporcionados por los microt&uacute;bulos del huso de anafase (MTs), pero c&oacute;mo se generan esas se&ntilde;ales es incierta. Usando qu&iacute;mica gen&eacute;tica para alcanzar control espacial y temporal, demostramos que la entrega autoorganizada de Polo-como quinasa 1 (Plk1) a la midzone y su fosforilaci&oacute;n local de un substrato de MT-limite son cr&iacute;ticos para la generaci&oacute;n de esta se&ntilde;al inductora de surco. Cuando Plk1 fue activa pero no puede dirigirse a s&iacute; mismo a este monumento Ecuatorial, ambas corticales inducci&oacute;n de reclutamiento y surco de RhoA no ocurren, as&iacute; recapitulando los efectos de inhibici&oacute;n de Plk1 anafase espec&iacute;ficos. Mediante espectrometr&iacute;a de masas en t&aacute;ndem y anticuerpos phosphospecific, encontramos que Plk1 se une y fosforila directamente la subunidad HsCYK-4 de centralspindlin (tambi&eacute;n conocido como MgcRacGAP) en el midzone. En serina 157, esta modificaci&oacute;n crea un importante sitio de acoplamiento para el t&aacute;ndem que BRCT repite el factor de intercambio de GTP Rho Ect2. Las c&eacute;lulas que expresan s&oacute;lo una forma de nonphosphorylatable de HsCYK-4 se ha podido localizar Ect2 en el midzone y fueron severamente deterioradas en la formaci&oacute;n del surco escote, lo que implica que HsCYK-4 es tarifa-limitaci&oacute;n blanco de Plk1 aguas arriba de RhoA. Por el contrario, atar a un alelo resistente al inhibidor de Plk1 a HsCYK-4 permitida surcos para formar a pesar de la inhibici&oacute;n global de todas otras mol&eacute;culas de Plk1 en la c&eacute;lula. Nuestros resultados iluminan dos mecanismos fundamentales que rigen la iniciaci&oacute;n de la citocinesis en c&eacute;lulas humanas e ilustran el poder de la qu&iacute;mica gen&eacute;tica para sondear esa reglamentaci&oacute;n tanto en tiempo y espacio.

Fosforilaci&oacute;n de cebado de HsCYK-4 en el midzone del huso y autoorganizaci&oacute;n de Plk1 regulan el inicio de la divisi&oacute;n de las c&eacute;lulas humanas.

Acquired resistance to Docetaxel precedes fatality in hormone-refractory prostate cancer (HRPC). However, strategies that target Docetaxel resistant cells remain elusive. Using in vitro and in vivo models, we identified a subpopulation of cells that survive Docetaxel exposure. This subpopulation lacks differentiation markers and HLA class I (HLAI) antigens, while overexpressing the Notch and Hedgehog signaling pathways. These cells were found in prostate cancer tissues and were related to tumor aggressiveness and poor patient prognosis. Notably, targeting Notch and Hedgehog signaling depleted this population through inhibition of the survival molecules AKT and Bcl-2, suggesting a therapeutic strategy for abrogating Docetaxel resistance in HRPC. Finally, these cells exhibited potent tumor-initiating capacity, establishing a link between chemotherapy resistance and tumor progression.

Suppression of acquired docetaxel resistance in prostate cancer through depletion of notch- and hedgehog-dependent tumor-initiating cells.

The spindle assembly checkpoint (SAC) delays anaphase until all chromosomes are bioriented on the mitotic spindle. Under current models, unattached kinetochores transduce the SAC by catalyzing the intramitotic production of a diffusible inhibitor of APC/C(Cdc20) (the anaphase-promoting complex/cyclosome and its coactivator Cdc20, a large ubiquitin ligase). Here we show that nuclear pore complexes (NPCs) in interphase cells also function as scaffolds for anaphase-inhibitory signaling. This role is mediated by Mad1-Mad2 complexes tethered to the nuclear basket, which activate soluble Mad2 as a binding partner and inhibitor of Cdc20 in the cytoplasm. Displacing Mad1-Mad2 from nuclear pores accelerated anaphase onset, prevented effective correction of merotelic errors, and increased the threshold of kinetochore-dependent signaling needed to halt mitosis in response to spindle poisons. A heterologous Mad1-NPC tether restored Cdc20 inhibitor production and normal M phase control. We conclude that nuclear pores and kinetochores both emit "wait anaphase" signals that preserve genome integrity.

Nuclear pores protect genome integrity by assembling a premitotic and mad1-dependent anaphase inhibitor.

Elucidating the determinants of aggressiveness in lethal prostate cancer may stimulate therapeutic strategies that improve clinical outcomes. We used experimental models and clinical databases to identify GATA2 as a regulator of chemotherapy resistance and tumorigenicity in this context. Mechanistically, direct upregulation of the growth hormone IGF2 emerged as a mediator of the aggressive properties regulated by GATA2. IGF2 in turn activated IGF1R and INSR as well as a downstream polykinase program. The characterization of this axis prompted a combination strategy whereby dual IGF1R/INSR inhibition restored the efficacy of chemotherapy and improved survival in preclinical models. These studies reveal a GATA2-IGF2 aggressiveness axis in lethal prostate cancer and identify a therapeutic opportunity in this challenging disease.

A targetable GATA2-IGF2 axis confers aggressiveness in lethal prostate cancer.

Advanced prostate cancer is a classic example of the intractability and consequent lethality that characterizes metastatic carcinomas. Novel treatments have improved the survival of men with prostate cancer; however, advanced prostate cancer invariably becomes resistant to these therapies and ultimately progresses to a lethal metastatic stage. Consequently, detailed knowledge of the molecular mechanisms that control prostate cancer cell survival and progression towards this lethal stage of disease will benefit the development of new therapeutics. The transcription factor endothelial transcription factor GATA-2 (GATA2) has been reported to have a key role in driving prostate cancer aggressiveness. In addition to being a pioneer transcription factor that increases androgen receptor (AR) binding and activity, GATA2 regulates a core subset of clinically relevant genes in an AR-independent manner. Functionally, GATA2 overexpression in prostate cancer increases cellular motility and invasiveness, proliferation, tumorigenicity, and resistance to standard therapies. Thus, GATA2 has a multifaceted function in prostate cancer aggressiveness and is a highly attractive target in the development of novel treatments against lethal prostate cancer.

The role of GATA2 in lethal prostate cancer aggressiveness.

PP2A comprising B56 regulatory subunit isoforms (PP2A(B56)) is a serine/threonine phosphatase essential for mitosis. At the kinetochore, PP2A(B56) both stabilizes microtubule binding and promotes silencing of the spindle assembly checkpoint (SAC) through its association with the SAC protein BubR1. Cells depleted of the B56 regulatory subunits of PP2A are delayed in activation of Cdc20-containing APC/C (APC/C(Cdc20)), which is an essential step for mitotic exit. It has been hypothesized that this delay arises from increased production of the mitotic checkpoint complex (MCC), an APC/C(Cdc20) inhibitor formed at unattached kinetochores through SAC signaling. In contrast to this prediction, we show that depletion of B56 subunits does not increase the amount or stability of the MCC. Rather, delays in APC/C(Cdc20) activation in B56-depleted cells correlate with impaired Cdc20 binding to APC/C. Stimulation of APC/C(Cdc20) assembly does not require binding between PP2A(B56) and BubR1, and thus this contribution of PP2A(B56) towards mitotic exit is distinct from its functions at kinetochores. PP2A(B56) associates with APC/C constitutively in a BubR1-independent manner. A mitotic phosphorylation site on Cdc20, known to be a substrate of PP2A(B56), modulates APC/C(Cdc20) assembly. These results elucidate the contributions of PP2A(B56) towards completion of mitosis.

The PP2A(B56) phosphatase promotes the association of Cdc20 with APC/C in mitosis.

Human leukocyte antigen class I (HLA-I) down-regulation has been reported in many human cancers to be associated with poor clinical outcome. However, its connection to tumor-initiating cells (TICs) remains unknown. In this study, we report that HLA-I is down-regulated in a subpopulation of cells that have high tumor initiating capacity in different types of human sarcomas. Detailed characterization revealed their distinct molecular profiles regarding proliferation, apoptosis and stemness programs. Notably, these TICs can be induced to differentiate along distinct mesenchymal lineages, including the osteogenic pathway. The retinoic acid receptor signaling pathway is overexpressed in HLA-1 negative TICs. All-trans retinoic acid treatment successfully induced osteogenic differentiation of this subpopulation, in vitro and in vivo, resulting in significantly decreased tumor formation. Thus, our findings indicate down-regulated HLA-I is a shared feature of TICs in a variety of human sarcomas, and differentiation therapy strategies may specifically target undifferentiated TICs and inhibit tumor formation.

Targeting sarcoma tumor-initiating cells through differentiation therapy.

The spindle assembly checkpoint kinase Mps1 not only inhibits anaphase but also corrects erroneous attachments that could lead to missegregation and aneuploidy. However, Mps1's error correction-relevant substrates are unknown. Using a chemically tuned kinetochore-targeting assay, we show that Mps1 destabilizes microtubule attachments (K fibers) epistatically to Aurora B, the other major error-correcting kinase. Through quantitative proteomics, we identify multiple sites of Mps1-regulated phosphorylation at the outer kinetochore. Substrate modification was microtubule sensitive and opposed by PP2A-B56 phosphatases that stabilize chromosome-spindle attachment. Consistently, Mps1 inhibition rescued K-fiber stability after depleting PP2A-B56. We also identify the Ska complex as a key effector of Mps1 at the kinetochore-microtubule interface, as mutations that mimic constitutive phosphorylation destabilized K fibers in vivo and reduced the efficiency of the Ska complex's conversion from lattice diffusion to end-coupled microtubule binding in vitro. Our results reveal how Mps1 dynamically modifies kinetochores to correct improper attachments and ensure faithful chromosome segregation.

Mps1 Regulates Kinetochore-Microtubule Attachment Stability via the Ska Complex to Ensure Error-Free Chromosome Segregation.

Nuclear pore complexes (NPCs) regulate nuclear-cytoplasmic transport, transcription, and genome integrity in eukaryotic cells. However, their functional roles in cancer remain poorly understood. We interrogated the evolutionary transcriptomic landscape of NPC components, nucleoporins (Nups), from primary to advanced metastatic human prostate cancer (PC). Focused loss-of-function genetic screen of top-upregulated Nups in aggressive PC models identified POM121 as a key contributor to PC aggressiveness. Mechanistically, POM121 promoted PC progression by enhancing importin-dependent nuclear transport of key oncogenic (E2F1, MYC) and PC-specific (AR-GATA2) transcription factors, uncovering a pharmacologically targetable axis that, when inhibited, decreased tumor growth, restored standard therapy efficacy, and improved survival in patient-derived pre-clinical models. Our studies molecularly establish a role of NPCs in PC progression and give a rationale for NPC-regulated nuclear import targeting as a therapeutic strategy for lethal PC. These findings may have implications for understanding how NPC deregulation contributes to the pathogenesis of other tumor types.

ArticleTotal : 4	Year
Publication title	2014
Publication title	2015
Publication title	2017
Publication title	2019

Article	Year
Chk1- and claspin-dependent but ATR/ATM- and Rad17-independent DNA replication checkpoint response in HeLa cells. Cancer research\| PubMed ID: 16951182	2006
Different S/M checkpoint responses of tumor and non tumor cell lines to DNA replication inhibition. Cancer research\| PubMed ID: 18089794	2007
The decision to enter mitosis: feedback and redundancy in the mitotic entry network. The Journal of cell biology\| PubMed ID: 19364923	2009
Plk1 self-organization and priming phosphorylation of HsCYK-4 at the spindle midzone regulate the onset of division in human cells. PLoS biology\| PubMed ID: 19468302	2009
Suppression of acquired docetaxel resistance in prostate cancer through depletion of notch- and hedgehog-dependent tumor-initiating cells. Cancer cell\| PubMed ID: 22975379	2012
Nuclear pores protect genome integrity by assembling a premitotic and mad1-dependent anaphase inhibitor. Cell\| PubMed ID: 24581499	2014
A targetable GATA2-IGF2 axis confers aggressiveness in lethal prostate cancer. Cancer cell\| PubMed ID: 25670080	2015
The role of GATA2 in lethal prostate cancer aggressiveness. Nature reviews. Urology\| PubMed ID: 27872477	2017
The PP2A(B56) phosphatase promotes the association of Cdc20 with APC/C in mitosis. Journal of cell science\| PubMed ID: 28404789	2017
Targeting sarcoma tumor-initiating cells through differentiation therapy. Stem cell research\| PubMed ID: 28433655	2017
Mps1 Regulates Kinetochore-Microtubule Attachment Stability via the Ska Complex to Ensure Error-Free Chromosome Segregation. Developmental cell\| PubMed ID: 28441529	2017
Nuclear Pores Promote Lethal Prostate Cancer by Increasing POM121-Driven E2F1, MYC, and AR Nuclear Import. Cell\| PubMed ID: 30100187	2018

Veronica Rodriguez-Bravo

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