The <em>Drosophila</em> Imaginal Disc Tumor Model: Visualization and Quantification of Gene Expression and Tumor Invasiveness Using Genetic Mosaics

Drosophila melanogaster has emerged as a powerful experimental system for functional and mechanistic studies of tumor development and progression in the ...

University of Cologne

Germline transformation with new transposon vectors now enables causal tests of gene function via ectopic protein expression or RNA interference in non-drosophilid insects. The problem remains of how to drive the transgene expression in vivo. We employed germline transformation using the piggyBac 3xP3-EGFP vector to test whether the Drosophila heat shock hsp70 promoter will be active in the live silkworm. We modified the original vector by cloning the coding sequence for Bombyx nuclear receptor Ftz-F1 between the hsp70 promoter and the terminator. Three independent transgenic lines expressing the Pax-6-driven EGFP marker in larval and adult photoreceptors were obtained with efficiencies of up to 1.7% of fertile G0 adults that gave GFP-positive progeny. Chromosomal integration of the transposon was confirmed with inverse PCR. Heat induction of the transgenic BmFtz-F1 was proven at both the mRNA and protein levels. RT-PCR data showed that the Drosophila heat shock promoter was functional in all three transgenic lines. Although basal activity was apparent at 25 degrees C, 1 h at 42 degrees C induced BmFtz-F1 mRNA at different stages of development and in diverse tissues. The relative levels of induction differed among the transgenic lines. Northern blot hybridization detected transgenic BmFtz-F1 only after heat shock and low levels of the mRNA were still present 6 h after the heat treatment. Immunostaining of epidermis using anti-BmFtz-F1 antibody showed a clear increase of nuclear signal 90 min after a heat shock.

Heat-inducible transgenic expression in the silkmoth Bombyx mori.

&#x8F6C;&#x5F55;&#x56E0;&#x5B50;&#x5206;&#x5316;&#x7684;&#x6210;&#x4EBA;&#x7ED3;&#x6784;&#x4EE5;&#x53CA;&#x5BF9;&#x4E8E;&#x8FC7;&#x65F6;&#x7684;&#x5E7C;&#x866B;&#x673A;&#x5173;&#x679C;&#x8747;&#x679C;&#x8747;&#x53D8;&#x6001;&#x8FC7;&#x7A0B;&#x7F16;&#x7A0B;&#x6027;&#x6B7B;&#x4EA1;&#x9700;&#x8981;&#x5E7F;&#x6CDB;&#x590D;&#x6742; (BR-C)&#x3002;&#x4E0D;&#x53EF;&#x4EE5;&#x65E0;&#x635F;&#x5931;&#x7684;&#x529F;&#x80FD;&#x57FA;&#x56E0;&#x6D4B;&#x8BD5;&#xFF0C;&#x5728;&#x975E; drosophilid &#x7269;&#x79CD;&#x4E2D;&#x6267;&#x884C;&#x8BC1;&#x660E;&#x662F;&#x5426; BR C &#x5176;&#x4ED6; holometabolous &#x6606;&#x866B;&#x5177;&#x6709;&#x7C7B;&#x4F3C;&#x7684;&#x4F5C;&#x7528;&#x3002;&#x5728;&#x8FD9;&#x91CC;&#x6211;&#x4EEC;&#x4F7F;&#x7528;&#x91CD;&#x7EC4;&#x7684;&#x8F9B;&#x5FB7;&#x6BD5;&#x65AF;&#x75C5;&#x6BD2;&#x4F5C;&#x4E3A;&#x4E00;&#x79CD;&#x5DE5;&#x5177;&#x6765;&#x6C89;&#x9ED8;&#x4E0E;&#x5C55;&#x671B;&#x5BB6;&#x8695; BR C &#x8868;&#x8FBE;&#x5F0F;&#x3002;&#x8868;&#x793A;&#x4E00;&#x4E2A; BR C &#x53CD;&#x4E49; RNA &#x7247;&#x6BB5;&#x7684;&#x75C5;&#x6BD2;&#x51CF;&#x5C11;&#x5185;&#x6E90;&#x6027;&#x611F;&#x67D3;&#x7EC4;&#x7EC7; (&#x6210;&#x4EBA;&#x7FFC;&#x548C;&#x817F;&#x539F;&#x57FA;) &#x901A;&#x8FC7; RNA &#x5E72;&#x6270; (RNAi) BR C mRNA &#x6C34;&#x5E73;&#x3002;BR C RNAi &#x649E;&#x5BFC;&#x81F4;&#x5931;&#x8D25;&#x7684;&#x52A8;&#x7269;&#x5B8C;&#x6210;&#x5E7C;&#x866B;&#x86F9;&#x7684;&#x8FC7;&#x6E21;&#x4E2D;&#x6216;&#x5728;&#x7A0D;&#x540E;&#x7684;&#x5F62;&#x6001;&#x53D1;&#x751F;&#x7F3A;&#x9677;&#xFF0C;&#x5305;&#x62EC;&#x6210;&#x4EBA;&#x590D;&#x773C;&#x3001; &#x817F;&#x548C;&#x7FC5;&#x8180;&#x6765;&#x81EA;&#x5176;&#x5E7C;&#x866B;&#x7684;&#x7956;&#x7EC6;&#x80DE;&#x7684;&#x5206;&#x5316;&#x3002;BR C RNAi &#x8FD8;&#x6444;&#x52A8;&#x5E7C;&#x866B;&#x4E1D;&#x817A;&#x7684;&#x7EC6;&#x80DE;&#x7A0B;&#x5E8F;&#x6027;&#x7684;&#x7684;&#x6B7B;&#x4EA1;&#x3002;&#x8FD9;&#x4E9B;&#x53D1;&#x80B2;&#x7F3A;&#x9677;&#x5BF9;&#x5E94;&#x635F;&#x5931;&#x7684;&#x51FD;&#x6570;&#x8868;&#x578B;&#x7684; BR C &#x679C;&#x8747;&#x7A81;&#x53D8;&#x4F53;&#x7684;&#x5F62;&#x6001;&#x53D1;&#x751F;&#x548C;&#x9000;&#x884C;&#x6027;&#x65B9;&#x9762;&#x6697;&#x793A; BR C &#x5728;&#x8715;&#x53D8;&#x4E2D;&#x7684;&#x5173;&#x952E;&#x4F5C;&#x7528;&#x8FDB;&#x5316;&#x7684;&#x5B88;&#x6052;&#x3002;&#x6211;&#x4EEC;&#x4E5F;&#x8BC1;&#x660E;&#x8F9B;&#x5FB7;&#x6BD5;&#x65AF;&#x75C5;&#x6BD2;&#x662F;&#x65B0;&#x6606;&#x866B;&#x6A21;&#x578B;&#x4E2D;&#x53D1;&#x80B2;&#x57FA;&#x56E0;&#x7684;&#x6C89;&#x9ED8;&#x7684;&#x6709;&#x7528;&#x5DE5;&#x5177;&#x3002;

&#x4F7F;&#x7528;&#x7684;&#x8F9B;&#x5FB7;&#x6BD5;&#x65AF;&#x75C5;&#x6BD2;&#x4ECB;&#x5BFC; RNA &#x5E72;&#x6270;&#x8868;&#x73B0;&#x51FA;&#x5E7F;&#x6CDB;&#x590D;&#x6742;&#x5B88;&#x6052;&#x7684;&#x4F5C;&#x7528;&#x5728;&#x6606;&#x866B;&#x7684;&#x53D8;&#x6001;&#x3002;

The transcription factor Broad-Complex (BR-C) is required for differentiation of adult structures as well as for the programmed death of obsolete larval organs during metamorphosis of the fruit fly Drosophila melanogaster. Whether BR-C has a similar role in other holometabolous insects could not be proven without a loss-of-function genetic test, performed in a non-drosophilid species. Here we use a recombinant Sindbis virus as a tool to silence BR-C expression in the silkmoth Bombyx mori. The virus expressing a BR-C antisense RNA fragment reduced endogenous BR-C mRNA levels in infected tissues (adult wing and leg primordia) via RNA interference (RNAi). The RNAi knock-down of BR-C resulted in the failure of animals to complete the larval-pupal transition or in later morphogenetic defects, including differentiation of adult compound eyes, legs, and wings from their larval progenitors. BR-C RNAi also perturbed the programmed cell death of larval silk glands. These developmental defects correspond to loss-of-function phenotypes of BR-C Drosophila mutants in both the morphogenetic and degenerative aspects, suggesting that the critical role of BR-C in metamorphosis is evolutionarily conserved. We also demonstrate that the Sindbis virus is a useful vehicle for silencing of developmental genes in new insect models.

Use of Sindbis virus-mediated RNA interference to demonstrate a conserved role of Broad-Complex in insect metamorphosis.

Le facteur de transcription Broad-Complex (BR-C) est n&#xE9;cessaire pour la diff&#xE9;renciation des structures pour adultes ainsi que pour la mort programm&#xE9;e de la obsol&#xE8;tes organes larvaires pendant la m&#xE9;tamorphose de la mouche du vinaigre Drosophila melanogaster. Que BR-C joue un r&#xF4;le similaire dans d&#x26;#39;autres insectes holom&#xE9;taboles ne pouvait pas &#xEA;tre prouv&#xE9; sans un test de perte de fonction g&#xE9;n&#xE9;tique, r&#xE9;alis&#xE9;e dans une esp&#xE8;ce non-drosophilid. Ici, nous utilisons un virus Sindbis recombinant comme un outil pour faire taire BR-C d&#x26;#39;expression dans le mori Bombyx &#xE0; soie. Le virus exprimant un BR-C ARN antisens fragment r&#xE9;duit les niveaux d&#x26;#39;ARNm endog&#xE8;nes BR-C dans les tissus infect&#xE9;s (aile des adultes et &#xE9;bauches jambe) par l&#x26;#39;interm&#xE9;diaire d&#x26;#39;ARN interf&#xE9;rence (ARNi). Le RNAi knock-down de la BR-C a entra&#xEE;n&#xE9; l&#x26;#39;&#xE9;chec des animaux pour achever la transition des larves-nymphe ou des d&#xE9;fauts morphog&#xE9;n&#xE9;tiques tard, y compris la diff&#xE9;renciation des yeux compos&#xE9;s des adultes, des jambes, et les ailes de leurs anc&#xEA;tres larvaires. BR-C ARNi &#xE9;galement perturb&#xE9; la mort cellulaire programm&#xE9;e des glandes de soie des larves. Ces d&#xE9;fauts de d&#xE9;veloppement correspond &#xE0; la perte de fonction des ph&#xE9;notypes de mutants de drosophile BR-C, tant dans le morphog&#xE9;n&#xE9;tique et les aspects d&#xE9;g&#xE9;n&#xE9;ratives, ce qui sugg&#xE8;re que le r&#xF4;le critique de BR-C dans la m&#xE9;tamorphose est &#xE9;volutif conserv&#xE9;e. Nous d&#xE9;montrons &#xE9;galement que le virus Sindbis est un instrument utile pour faire taire des g&#xE8;nes du d&#xE9;veloppement dans de nouveaux mod&#xE8;les d&#x26;#39;insectes.

Utilisation de virus Sindbis m&#xE9;diation interf&#xE9;rence ARN pour d&#xE9;montrer le r&#xF4;le conserv&#xE9; de Broad-Complex dans La M&#xE9;tamorphose des insectes

Verwenden des Sindbis-Virus-vermittelte RNA-Interferenz, um eine konservierte Rolle von Broad-Complex in Insekten-Metamorphose zu demonstrieren.

Il fattore di trascrizione Broad-complesso (BR-C) &egrave; necessario per la differenziazione delle strutture adulti pure per quanto riguarda la morte programmata degli obsoleti organi larvali durante la metamorfosi del moscerino della frutta Drosophila melanogaster. Se BR-C ha un ruolo simile in altri insetti olometaboli non potrebbe essere dimostrata senza un test genetico di perdita di funzione, eseguito in una specie di non-drosophilid. Qui usiamo Sindbis virus ricombinanti come uno strumento per mettere a tacere i mori del bombice dell&#39;ailanto espressione BR-C. Il virus che esprimono un frammento di RNA antisenso BR-C ha ridotto i livelli di mRNA di BR-C endogeno nei tessuti infetti (adulti ala e gamba primordia) tramite interferenza del RNA (RNAi). Il RNAi knock-down del BR-C ha provocato il fallimento di animali per completare la transizione allo stadio larvale-pupa o in pi&ugrave; tardi morfogenetici difetti, tra cui differenziazione degli adulti gli occhi composti, le gambe e ali da loro progenitori larvale. BR-C RNAi turbare anche la morte programmata delle cellule delle ghiandole di seta larvale. Questi difetti di sviluppo corrispondono alla perdita di funzione fenotipi mutanti Drosophila BR-C entrambi gli aspetti morfogenetici e degenerative, suggerendo che il ruolo critico di BR-C in metamorfosi &egrave; evolutivamente conservato. Abbiamo anche dimostrare che il virus di Sindbis &egrave; un utile veicolo per il silenziamento di geni dello sviluppo di nuovi modelli di insetti.

Uso di interferenza del RNA mediata da virus Sindbis a dimostrare un ruolo conservato del vasto complesso in metamorfosi dell&#39;insetto.

&#x8EE2;&#x5199;&#x56E0;&#x5B50;&#xFF08;BR-C&#xFF09;&#x30D6;&#x30ED;&#x30FC;&#x30C9;&#xB7;&#x30B3;&#x30F3;&#x30D7;&#x30EC;&#x30C3;&#x30AF;&#x30B9;&#x306F;&#x3001;&#x5927;&#x4EBA;&#x306E;&#x69CB;&#x9020;&#x306E;&#x5206;&#x5316;&#x3068;&#x540C;&#x69D8;&#x306B;&#x30B7;&#x30E7;&#x30A6;&#x30B8;&#x30E7;&#x30A6;&#x30D0;&#x30A8;&#x306E;&#x5909;&#x614B;&#x4E2D;&#x306B;&#x5EC3;&#x6B62;&#x3055;&#x308C;&#x305F;&#x5E7C;&#x866B;&#x306E;&#x81D3;&#x5668;&#x306E;&#x30D7;&#x30ED;&#x30B0;&#x30E9;&#x30E0;&#x6B7B;&#x306E;&#x305F;&#x3081;&#x306B;&#x5FC5;&#x8981;&#x3068;&#x306A;&#x308A;&#x307E;&#x3059;&#x3002; BR-C&#x306F;&#x3001;&#x4ED6;&#x306E;&#x5B8C;&#x5168;&#x5909;&#x614B;&#x306E;&#x6606;&#x866B;&#x3067;&#x540C;&#x69D8;&#x306E;&#x5F79;&#x5272;&#x3092;&#x6301;&#x3063;&#x3066;&#x3044;&#x308B;&#x304B;&#x3069;&#x3046;&#x304B;&#x3092;&#x975E;&#x30B7;&#x30E7;&#x30A6;&#x30B8;&#x30E7;&#x30A6;&#x30D0;&#x30A8;&#x7A2E;&#x3067;&#x884C;&#x308F;&#x6A5F;&#x80FD;&#x55AA;&#x5931;&#x907A;&#x4F1D;&#x5B50;&#x691C;&#x67FB;&#x306A;&#x3057;&#x3067;&#x8A3C;&#x660E;&#x3055;&#x308C;&#x307E;&#x305B;&#x3093;&#x3067;&#x3057;&#x305F;&#x3002;&#x3053;&#x3053;&#x3067;&#x306F;&#x3001;&#x30AB;&#x30A4;&#x30B3;&#x30AC;&#x30AB;&#x30A4;&#x30B3;&#x306E;BR-C&#x306E;&#x767A;&#x73FE;&#x3092;&#x6291;&#x5236;&#x3059;&#x308B;&#x305F;&#x3081;&#x306E;&#x30C4;&#x30FC;&#x30EB;&#x3068;&#x200B;&#x200B;&#x3057;&#x3066;&#x3001;&#x7D44;&#x63DB;&#x3048;&#x30B7;&#x30F3;&#x30C9;&#x30D3;&#x30B9;&#x30A6;&#x30A4;&#x30EB;&#x30B9;&#x3092;&#x4F7F;&#x7528;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002; BR-C&#x30A2;&#x30F3;&#x30C1;&#x30BB;&#x30F3;&#x30B9;RNA&#x65AD;&#x7247;&#x3092;&#x767A;&#x73FE;&#x3059;&#x308B;&#x30A6;&#x30A4;&#x30EB;&#x30B9;&#x306F;&#x3001;RNA&#x5E72;&#x6E09;&#xFF08;RNAi&#xFF09;&#x3092;&#x4ECB;&#x3057;&#x3066;&#xFF08;&#x5927;&#x4EBA;&#x306E;&#x7FFC;&#x3068;&#x811A;&#x539F;&#x57FA;&#xFF09;&#x611F;&#x67D3;&#x7D44;&#x7E54;&#x306B;&#x304A;&#x3051;&#x308B;&#x5185;&#x56E0;&#x6027;BR-C mRNA&#x30EC;&#x30D9;&#x30EB;&#x3092;&#x6E1B;&#x5C11;&#x3055;&#x305B;&#x305F;&#x3002; BR-C&#x306E;RNAi&#x30CE;&#x30C3;&#x30AF;&#x30C0;&#x30A6;&#x30F3;&#x306F;&#x3001;&#x5E7C;&#x866B;&#xB7;&#x86F9;&#x306E;&#x79FB;&#x884C;&#x3092;&#x5B8C;&#x4E86;&#x3059;&#x308B;&#x306B;&#x306F;&#x3001;&#x52D5;&#x7269;&#x306E;&#x5931;&#x6557;&#x3067;&#x3001;&#x5927;&#x4EBA;&#x306E;&#x8907;&#x773C;&#x3001;&#x811A;&#x3001;&#x305D;&#x306E;&#x5E7C;&#x866B;&#x306E;&#x524D;&#x99C6;&#x7D30;&#x80DE;&#x304B;&#x3089;&#x306E;&#x7FFC;&#x306E;&#x5206;&#x5316;&#x3092;&#x542B;&#x3081;&#x305D;&#x308C;&#x4EE5;&#x964D;&#x306E;&#x5F62;&#x614B;&#x5F62;&#x6210;&#x306E;&#x6B20;&#x9665;&#x306B;&#x306A;&#x308A;&#x307E;&#x3057;&#x305F;&#x3002; BR-C RNAi&#x306F;&#x307E;&#x305F;&#x3001;&#x5E7C;&#x866B;&#x306E;&#x7D79;&#x7CF8;&#x817A;&#x306E;&#x30D7;&#x30ED;&#x30B0;&#x30E9;&#x30E0;&#x7D30;&#x80DE;&#x6B7B;&#x3092;&#x6442;&#x52D5;&#x3002;&#x3053;&#x308C;&#x3089;&#x306E;&#x767A;&#x9054;&#x969C;&#x5BB3;&#x306F;&#x3001;&#x5909;&#x614B;&#x4E2D;&#x306E;BR-C&#x306E;&#x91CD;&#x8981;&#x306A;&#x5F79;&#x5272;&#x304C;&#x9032;&#x5316;&#x7684;&#x306B;&#x4FDD;&#x5B58;&#x3055;&#x308C;&#x3066;&#x3044;&#x308B;&#x3053;&#x3068;&#x3092;&#x793A;&#x5506;&#x3057;&#x3001;&#x5F62;&#x614B;&#x5F62;&#x6210;&#x3084;&#x5909;&#x6027;&#x306E;&#x4E21;&#x9762;&#x3067;BR-C&#x30B7;&#x30E7;&#x30A6;&#x30B8;&#x30E7;&#x30A6;&#x30D0;&#x30A8;&#x306E;&#x5909;&#x7570;&#x4F53;&#x306E;&#x6A5F;&#x80FD;&#x55AA;&#x5931;&#x8868;&#x73FE;&#x578B;&#x306B;&#x5BFE;&#x5FDC;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x307E;&#x305F;&#x3001;&#x30B7;&#x30F3;&#x30C9;&#x30D3;&#x30B9;&#x30A6;&#x30A4;&#x30EB;&#x30B9;&#x306F;&#x3001;&#x65B0;&#x3057;&#x3044;&#x6606;&#x866B;&#x30E2;&#x30C7;&#x30EB;&#x306E;&#x767A;&#x9054;&#x306E;&#x907A;&#x4F1D;&#x5B50;&#x306E;&#x30B5;&#x30A4;&#x30EC;&#x30F3;&#x30B7;&#x30F3;&#x30B0;&#x306B;&#x6709;&#x7528;&#x306A;&#x624B;&#x6BB5;&#x3067;&#x3042;&#x308B;&#x3053;&#x3068;&#x3092;&#x793A;&#x3057;&#x3066;&#x3044;&#x308B;&#x3002;

&#x6606;&#x866B;&#x306E;&#x5909;&#x614B;&#x306B;&#x30D6;&#x30ED;&#x30FC;&#x30C9;&#x30B3;&#x30F3;&#x30D7;&#x30EC;&#x30C3;&#x30AF;&#x30B9;&#x306E;&#x4FDD;&#x5B58;&#x306E;&#x5F79;&#x5272;&#x3092;&#x5B9F;&#x8A3C;&#x3059;&#x308B;&#x30B7;&#x30F3;&#x30C9;&#x30D3;&#x30B9;&#x30A6;&#x30A4;&#x30EB;&#x30B9;&#x5A92;&#x4ECB;&#x6027;RNA&#x5E72;&#x6E09;&#x306E;&#x4F7F;&#x7528;

&#xB179;&#xC74C; &#xBC29;&#xC1A1; &#xC694;&#xC778; &#xAD11;&#xBC94;&#xC704; &#xD55C; &#xBCF5;&#xD569; (BR-C)&#xB294; &#xACFC;&#xC77C; &#xD30C;&#xB9AC; Drosophila melanogaster&#xC758; &#xBCC0; &#xD0DC; &#xD558;&#xB294; &#xB3D9;&#xC548; &#xC624;&#xB798; &#xB41C; &#xC560;&#xBC8C;&#xB808; &#xC7A5;&#xAE30;&#xC758; &#xD504;&#xB85C;&#xADF8;&#xB798;&#xBC0D; &#xB41C; &#xC8FD;&#xC74C;&#xC5D0; &#xAD00;&#xD574;&#xC11C;&#xB294; &#xC131;&#xC778; &#xAD6C;&#xC870; &#xB610;&#xD55C; &#xCC28;&#xBCC4;&#xD654; &#xD544;&#xC694; &#xD569;&#xB2C8;&#xB2E4;. &#xBE44; drosophilid &#xC885;&#xC5D0;&#xC11C; &#xC218;&#xD589; &#xAE30;&#xB2A5; &#xC190;&#xC2E4; &#xC720;&#xC804;&#xC790; &#xAC80;&#xC0AC; &#xC5C6;&#xC774; BR-C&#xB294; &#xB2E4;&#xB978; holometabolous &#xACE4;&#xCDA9;&#xC5D0; &#xBE44;&#xC2B7;&#xD55C; &#xC5ED;&#xD560;&#xC744; &#xC5EC;&#xBD80;&#xB97C; &#xC785;&#xC99D; &#xD558;&#xC9C0; &#xC218; &#xC5C6;&#xC2B5;&#xB2C8;&#xB2E4;. &#xC5EC;&#xAE30; &#xC6B0;&#xB9AC; silkmoth &#xB204;&#xC5D0;&#xB098;&#xBC29;&#xC5D0; BR C &#xC2DD; &#xCE68;&#xBB35; &#xB3C4;&#xAD6C;&#xB85C; &#xC7AC;&#xC870;&#xD569; Sindbis &#xBC14;&#xC774;&#xB7EC;&#xC2A4;&#xB97C; &#xC0AC;&#xC6A9;. BR-C &#xC13C;&#xC2A4; RNA &#xB2E8;&#xD3B8;&#xC744; &#xD45C;&#xD604; &#xD558;&#xB294; &#xBC14;&#xC774;&#xB7EC;&#xC2A4;&#xB294; RNA &#xAC04;&#xC12D; (RNAi)&#xC744; &#xD1B5;&#xD574; &#xAC10;&#xC5FC; &#xB41C; &#xC870;&#xC9C1; (&#xC131;&#xC778; &#xB0A0;&#xAC1C;&#xC640; &#xB2E4;&#xB9AC; primordia)&#xC5D0; &#xC0DD; BR C mRNA &#xC218;&#xC900; &#xAC10;&#xC18C;. BR-C&#xC758; RNAi &#xB178;&#xD06C; &#xB2E4;&#xC6B4; &#xC560;&#xBC8C;&#xB808; &#xBC88;&#xB370;&#xAE30; &#xC804;&#xD658;&#xC744; &#xC644;&#xB8CC; &#xD558;&#xB294; &#xB3D9;&#xBB3C;&#xC758; &#xC2E4;&#xD328; &#xB610;&#xB294; &#xC131;&#xC778; &#xD654;&#xD569;&#xBB3C; &#xB208;, &#xB2E4;&#xB9AC;, &#xADF8;&#xB9AC;&#xACE0; &#xADF8;&#xB4E4;&#xC758; &#xC560;&#xBC8C;&#xB808; progenitors&#xC5D0;&#xC11C; &#xB0A0;&#xAC1C;&#xC758; &#xCC28;&#xBCC4;&#xD654;&#xB97C; &#xD3EC;&#xD568; &#xD55C; &#xC804;&#xCCB4;&#xC801; &#xACB0;&#xD568; &#xB098;&#xC911; &#xACB0;&#xACFC;. BR-C Rnai&#xB294; &#xB610;&#xD55C; &#xC560;&#xBC8C;&#xB808; &#xC2E4;&#xD06C; &#xB540; &#xC0D8;&#xC758; &#xD504;&#xB85C;&#xADF8;&#xB7A8; &#xB41C; &#xC138;&#xD3EC; &#xC8FD;&#xC74C; &#xBD88;&#xC548;&#xC815;. &#xC774;&#xB7EC;&#xD55C; &#xBC1C;&#xB2EC; &#xACB0;&#xD568; &#xBCC0;&#xD615;&#xC5D0; BR-C&#xC758; &#xC911;&#xC694; &#xD55C; &#xC5ED;&#xD560; &#xC9C4;&#xD654; &#xC801;&#xC73C;&#xB85C; &#xBCF4;&#xC874;&#xC740; &#xC81C;&#xC548; &#xC804;&#xCCB4;&#xC801; &#xBC0F; &#xD1F4;&#xD589; &#xC131; &#xCE21;&#xBA74;&#xC5D0;&#xC11C; BR C &#xCD08;&#xD30C;&#xB9AC; &#xB3CC;&#xC5F0;&#xBCC0;&#xC774;&#xC758; &#xAE30;&#xB2A5; &#xC190;&#xC2E4; &#xACE0;&#xAE30;&#xC5D0; &#xD574;&#xB2F9; &#xD569;&#xB2C8;&#xB2E4;. &#xC6B0;&#xB9AC;&#xB294; &#xB610;&#xD55C; Sindbis &#xBC14;&#xC774;&#xB7EC;&#xC2A4;&#xB294; &#xC0C8;&#xB85C;&#xC6B4; &#xACE4;&#xCDA9; &#xBAA8;&#xB378; &#xAC1C;&#xBC1C; &#xC720;&#xC804;&#xC790;&#xC758; &#xC785;&#xC744; &#xC704;&#xD55C; &#xC720;&#xC6A9;&#xD55C; &#xCC28;&#xB7C9; &#xBCF4;&#xC5EC; &#xC90D;&#xB2C8;&#xB2E4;.

&#xC0AC;&#xC6A9;&#xC758; &#xAD11;&#xBC94;&#xC704; &#xD55C; &#xB2E8;&#xC9C0;&#xC758; &#xBCF4;&#xC874;&#xB41C; &#xC5ED;&#xD560;&#xC744; &#xBCF4;&#xC5EC; &#xC8FC;&#xB294; Sindbis &#xBC14;&#xC774;&#xB7EC;&#xC2A4; &#xC911;&#xC7AC; RNA &#xAC04;&#xC12D;&#xC5D0; &#xACE4;&#xCDA9; &#xBCC0; &#xD0DC;.

El factor de transcripci&#xF3;n de amplio complejo (BR-C) es necesaria para la diferenciaci&#xF3;n de las estructuras adultas, as&#xED; como para la muerte programada de los &#xF3;rganos obsoletos larvas durante la metamorfosis de la mosca de la fruta Drosophila melanogaster. Si BR-C tiene un papel similar en otros insectos holomet&#xE1;bolos no se pod&#xED;a probar sin una prueba de la p&#xE9;rdida de la funci&#xF3;n gen&#xE9;tica, realizado en una especie no drosofilidos. Aqu&#xED; se utiliza un virus Sindbis recombinante como una herramienta para silenciar a la BR-C expresi&#xF3;n en el Bombyx mori silkmoth. El virus de la expresi&#xF3;n de una BR-C fragmento de ARN antisentido redujo end&#xF3;genos BR-C los niveles de mRNA en tejidos infectados (ala de los adultos y el primordio de la pierna) a trav&#xE9;s de ARN de interferencia (RNAi). El RNAi knock-down de la BR-C dio como resultado el fracaso de los animales para completar la transici&#xF3;n de larvas-pupas o en defectos morfogen&#xE9;ticos m&#xE1;s adelante, incluyendo la diferenciaci&#xF3;n de los ojos compuestos de adultos, las piernas y las alas de sus progenitores larvas. BR-C RNAi tambi&#xE9;n perturb&#xF3; la muerte programada de las c&#xE9;lulas de las gl&#xE1;ndulas de seda larvas. Estos defectos en el desarrollo corresponden a fenotipos de p&#xE9;rdida de la funci&#xF3;n de la BR-C mutantes de Drosophila, tanto en la morfog&#xE9;nesis y aspectos degenerativos, lo que sugiere que el papel fundamental de la BR-C en la metamorfosis est&#xE1; conservado evolutivamente. Tambi&#xE9;n demuestran que el virus Sindbis es un veh&#xED;culo &#xFA;til para el silenciamiento de genes del desarrollo de nuevos modelos de insectos.

El uso de Sindbis virus mediada por la interferencia de ARN para demostrar una funci&#xF3;n conservada del amplio complejo en la metamorfosis de los insectos

Basic leucine zipper proteins Jun and Fos form the dimeric transcription factor AP-1, essential for cell differentiation and immune and antioxidant defenses. AP-1 activity is controlled, in part, by the redox state of critical cysteine residues within the basic regions of Jun and Fos. Mutation of these cysteines contributes to oncogenic potential of Jun and Fos. How cells maintain the redox-dependent AP-1 activity at favorable levels is not known. We show that the conserved coactivator MBF1 is a positive modulator of AP-1. Via a direct interaction with the basic region of Drosophila Jun (D-Jun), MBF1 prevents an oxidative modification (S-cystenyl cystenylation) of the critical cysteine and stimulates AP-1 binding to DNA. Cytoplasmic MBF1 translocates to the nucleus together with a transfected D-Jun protein, suggesting that MBF1 protects nascent D-Jun also in Drosophila cells. mbf1-null mutants live shorter than mbf1+ controls in the presence of hydrogen peroxide (H2O2). An AP-1-dependent epithelial closure becomes sensitive to H2O2 in flies lacking MBF1. We conclude that by preserving the redox-sensitive AP-1 activity, MBF1 provides an advantage during oxidative stress.

Coactivator MBF1 preserves the redox-dependent AP-1 activity during oxidative stress in Drosophila.

The role of c-Jun N-terminal kinase (JNK) signaling in cancer is enigmatic, and both tumor-promoting and tumor-suppressing functions have been ascribed to JNK pathway components. We have used the Drosophila eye to investigate the function of the JNK pathway in three different tumor models of increasing malignancy. Benign lesions caused by loss of the neoplastic tumor suppressor gene scribble can efficiently be eliminated by JNK-induced apoptosis. In such a scenario, the eye reverts to a wild-type phenotype, indicating that the JNK pathway prevents tumor formation. The situation changes in the case of aggressive tissue overgrowth, which can be induced by oncogenic activation of the Ras/Raf pathway in the eye, or in malignant invasive tumors resulting when Raf activation is combined with loss of scribble. The growth of these more aggressive tumor types is significantly, yet incompletely, suppressed by JNK-mediated apoptosis. Remarkably, oncogenic Raf and JNK cooperate in these tumors, to induce massive hyperplasia in adjacent wild-type tissue. Thus, depending on the genetic context, JNK signaling can eradicate tumors by removing premalignant cells, or promote aberrant overgrowth in tissues surrounding primary lesions.

Non-cell-autonomous induction of tissue overgrowth by JNK/Ras cooperation in a Drosophila tumor model.

Le r&ocirc;le de la kinase c-Jun N-terminal (JNK) de signalisation dans le cancer est &eacute;nigmatique, et fonctions fois favorisant la tumeur et suppresseur de tumeur ont &eacute;t&eacute; attribu&eacute;es aux composants de voie JNK. Nous avons utilis&eacute; le &oelig;il de la drosophile pour &eacute;tudier la fonction de la voie JNK dans trois mod&egrave;les diff&eacute;rents de tumeur de plus en plus une tumeur maligne. Des l&eacute;sions b&eacute;nignes, caus&eacute;es par la perte de la scribble de g&egrave;ne suppresseur tumeur n&eacute;oplasiques peuvent efficacement &ecirc;tre &eacute;limin&eacute;es par l&#39;apoptose induite par JNK. Dans un tel sc&eacute;nario, le &oelig;il revient &agrave; un ph&eacute;notype sauvage, indiquant que la voie JNK emp&ecirc;che la formation de tumeurs. La situation change dans le cas de la prolif&eacute;ration de tissu agressif, qui peut &ecirc;tre induite par l&#39;oncog&egrave;ne activation de la voie Ras/Raf dans le &oelig;il, ou dans les tumeurs malignes envahissantes r&eacute;sultant lors de l&#39;activation de la Raf est associ&eacute;e &agrave; la perte de dessin &agrave; main lev&eacute;e. La croissance de ces types de tumeur plus agressives est significativement, mais de mani&egrave;re incompl&egrave;te, supprim&eacute;e par apoptose m&eacute;di&eacute;e par JNK. Remarquablement, les oncog&egrave;nes Raf et JNK coop&egrave;rent dans ces tumeurs, d&#39;induire une hyperplasie massive dans les tissus adjacents de type sauvage. Ainsi, selon le contexte g&eacute;n&eacute;tique, JNK signalisation peut &eacute;liminer des tumeurs en supprimant les cellules pr&eacute;canc&eacute;reuses ou promouvoir la prolif&eacute;ration aberrante dans les tissus entourant les l&eacute;sions primaires.

Non-cellule autonome induction de la surcroissance de tissus par JNK/Ras coop&eacute;ration dans un mod&egrave;le de tumeur de drosophile.

Il ruolo di c-Jun N-terminal chinasi (JNK) segnalazione nel cancro &egrave; enigmatico e funzioni sia tumore-promozione e soppressione del tumore sono state attribuite ai componenti del pathway di JNK. Abbiamo usato l&#39;occhio della drosofila per indagare la funzione del percorso in tre modelli diversi di tumore crescente malignit&agrave; JNK. Lesioni benigne causate da perdita dello scarabocchio di gene soppressore tumorale neoplastica possono essere eliminate in modo efficiente da apoptosi indotta da JNK. In tale scenario, l&#39;occhio viene ripristinato un fenotipo wild-type, che indica che il pathway JNK impedisce la formazione di tumori. La situazione cambia nel caso di crescita eccessiva di tessuto aggressivo, che pu&ograve; essere indotta da attivazione oncogenica della via Ras/Raf nell&#39;occhio, o in tumori maligni invasivi, con conseguente attivazione di Raf &egrave; combinati con perdita del disegno a mano libera. La crescita di questi tipi di tumore pi&ugrave; aggressivi &egrave; significativamente, ancora incompleto, soppressa dal apoptosis JNK-mediata. Notevolmente, oncogenica Raf e JNK cooperare in questi tumori, di indurre iperplasia massiccia nel selvaggio-tipo tessuti adiacenti. Cos&igrave;, a seconda del contesto genetico, JNK segnalazione pu&ograve; sradicare i tumori rimuovendo le cellule precancerose, o promuovere la proliferazione aberrante nei tessuti circostanti lesioni primarie.

Non-cella autonomi induzione della crescita eccessiva di tessuto dalla cooperazione di JNK/Ras in un modello di tumore della drosofila.

El papel de la quinasa c-Jun N-terminal (JNK) se&ntilde;alizaci&oacute;n en c&aacute;ncer es enigm&aacute;tico, y funciones de promoci&oacute;n de tumor tanto supresoras de tumor se han atribuido a los componentes de v&iacute;a JNK. Hemos utilizado el ojo de la Drosophila para investigar la funci&oacute;n de la v&iacute;a JNK en tres modelos diferentes de tumor de aumento de la malignidad. Lesiones benignas, causadas por la p&eacute;rdida del garabato de gen supresor de tumor neopl&aacute;sico se pueden eliminar eficientemente por apoptosis inducida por JNK. En tal escenario, el ojo vuelve a un fenotipo de tipo salvaje, lo que indica que la v&iacute;a JNK previene la formaci&oacute;n de tumores. La situaci&oacute;n cambia en el caso de crecimiento excesivo del tejido agresivo, que puede ser inducida por activaci&oacute;n oncog&eacute;nica de la v&iacute;a Ras/Raf en el ojo, o en los tumores malignos invasivos que resulta cuando la activaci&oacute;n de la Royal Air Force se combina con p&eacute;rdida de garabato. El crecimiento de este tipo de tumor m&aacute;s agresivo es significativamente, pero incompleto, suprimido por apoptosis mediado de JNK. Notable, oncog&eacute;nicos Raf y JNK cooperan en estos tumores, para inducir hiperplasia masiva en tejido adyacente de tipo salvaje. As&iacute;, dependiendo del contexto gen&eacute;tico, se&ntilde;alizaci&oacute;n JNK puede erradicar tumores mediante la eliminaci&oacute;n de las c&eacute;lulas premalignas o promover crecimiento aberrante excesivo en los tejidos que rodean las lesiones primarias.

Inducci&oacute;n no celular aut&oacute;nomo del crecimiento excesivo del tejido por JNK/Ras cooperaci&oacute;n en un modelo de tumor de Drosophila.

Perte de la polarit&eacute; &eacute;pith&eacute;liale g&egrave;ne scribble dans les clones de la drosophile, les cellules du disque imaginal peuvent coop&eacute;rer avec Ras signalisation d&#39;induire des tumeurs malignes. Ces tissus mutant overproliferates, r&eacute;siste &agrave; l&#39;apoptose, laisse sa place d&#39;origine et envahit les autres organes, causant finalement la l&eacute;talit&eacute;. Nous montrons que l&#39;augmentation des Jun N-terminal kinase (JNK) signalisation r&eacute;sultant de la perte de griffonnage favorise la circulation des cellules transform&eacute;es de sites secondaires. Cet effet n&eacute;cessite l&#39;activation transcriptionnelle Fos d&eacute;pendant d&#39;une matrice m&eacute;talloprot&eacute;ase g&egrave;ne mmp1 en aval de JNK. Expression de la pr&eacute;cipitation de Timp ou Mmp RNAi d&#39;inhibiteur de la Mmp supprime le pouvoir envahissant de cellule. La fonction proinvasive de la voie JNK est r&eacute;v&eacute;l&eacute;e dans un contexte de tumeur lorsqu&#39;active Ras signalisation emp&ecirc;che la r&eacute;ponse apoptotique de l&#39;activit&eacute; JNK tel qu&#39;il appara&icirc;t dans les cellules non transform&eacute;es. Selon ces r&eacute;sultats, nous pr&eacute;sentons un mod&egrave;le qui explique la coop&eacute;ration oncog&eacute;nique entre JNK et Ras et d&eacute;crit comment aberrante r&eacute;gulation de la survie des cellules, prolif&eacute;ration et mobilisation coop&egrave;rent pour inciter &agrave; la formation de tumeurs malignes.

JNK et Fos-r&eacute;glement&eacute;s Mmp1 expression coop&egrave;re avec Ras d&#39;induire des tumeurs invasives chez la drosophile.

Loss of the epithelial polarity gene scribble in clones of Drosophila imaginal disc cells can cooperate with Ras signaling to induce malignant tumors. Such mutant tissue overproliferates, resists apoptosis, leaves its place of origin and invades other organs, ultimately causing lethality. We show that increased Jun N-terminal kinase (JNK) signaling resulting from the loss of scribble promotes the movement of transformed cells to secondary sites. This effect requires Fos-dependent transcriptional activation of a matrix metalloprotease gene mmp1 downstream of JNK. Expression of the Mmp inhibitor Timp or Mmp RNAi knockdown suppresses cell invasiveness. The proinvasive function of the JNK pathway is revealed in a tumor context when active Ras signaling prevents the apoptotic response to JNK activity as it occurs in nontransformed cells. Based on these results, we present a model that explains the oncogenic cooperation between JNK and Ras, and describes how aberrant regulation of cell survival, proliferation and mobilization cooperate to incite malignant tumor formation.

JNK- and Fos-regulated Mmp1 expression cooperates with Ras to induce invasive tumors in Drosophila.

Perdita del polarit&agrave; epiteliali gene scarabocchio in cloni di cellule disco immaginale possono cooperare con Ras di segnalazione per indurre tumori maligni della drosofila. Tale tessuto mutante overproliferates, resiste all&#39;apoptosi, lascia il suo luogo di origine e invade altri organi, causando alla fine la letalit&agrave;. Mostriamo che aumentato Jun N-terminal chinasi (JNK) segnalazione risultanti dalla perdita di scribble promuove la circolazione delle cellule trasformate a siti secondari. Questo effetto richiede attivazione trascrizionale Fos-dipendente di una matrice metalloprotease gene mmp1 a valle di JNK. Espressione dell&#39;atterramento Mmp inibitore Timp o Mmp RNAi sopprime invasivit&agrave; delle cellule. La funzione proinvasive della via JNK &egrave; rivelata in un contesto di tumore quando attivo di segnalazione Ras impedisce la risposta apoptotica al attivit&agrave; JNK come si verifica nelle cellule nontransformed. Basato su questi risultati, vi presentiamo un modello che spiega la cooperazione oncogenica tra JNK e Ras e descrive come aberrante regolazione della sopravvivenza cellulare, proliferazione e mobilitazione collaborano per incitare alla formazione di tumori maligni.

JNK e Fos-regolate Mmp1 espressione collabora con Ras di indurre tumori invasivi in drosofila.

P&eacute;rdida de la polaridad epitelial gene garabato en clones de Drosophila las c&eacute;lulas tipo disco imaginal pueden cooperar con Ras se&ntilde;alizaci&oacute;n inducir tumores malignos. Dicho tejido mutante overproliferates, resiste la apoptosis, deja su lugar de origen e invade otros &oacute;rganos, en &uacute;ltima instancia, causando mortalidad. Mostramos que aument&oacute; Jun N-terminal quinasa (JNK) se&ntilde;alizaci&oacute;n resultantes de la p&eacute;rdida de garabato promueve el movimiento de las c&eacute;lulas transformadas sitios secundarios. Este efecto requiere activaci&oacute;n transcripcional dependiente de Fos de una matriz metaloproteinasa gene mmp1 aguas abajo de JNK. Expresi&oacute;n de la ca&iacute;da de inhibidor Timp o Mmp RNAi Mmp suprime invasividad celular. La funci&oacute;n proinvasive de la v&iacute;a JNK se revela en un contexto de tumor cuando activo Ras se&ntilde;alizaci&oacute;n previene la respuesta apopt&oacute;tica a la actividad JNK como ocurre en las c&eacute;lulas nontransformed. Basado en estos resultados, se presenta un modelo que explica la cooperaci&oacute;n oncog&eacute;nica entre JNK y Ras y describe c&oacute;mo aberrante Reglamento de supervivencia celular, proliferaci&oacute;n y movilizaci&oacute;n cooperan para incitar a la formaci&oacute;n de tumores malignos.

Regulado de JNK y Fos Mmp1 expresi&oacute;n coopera con Ras para inducir tumores invasivos en Drosophila.

Epithelial sheet spreading and fusion underlie important developmental processes. Well-characterized examples of such epithelial morphogenetic events have been provided by studies in Drosophila, and include embryonic dorsal closure, formation of the adult thorax and wound healing. All of these processes require the basic region-leucine zipper (bZIP) transcription factors Jun and Fos. Much less is known about morphogenesis of the fly abdomen, which involves replacement of larval epidermal cells (LECs) with adult histoblasts that divide, migrate and finally fuse to form the adult epidermis during metamorphosis. Here, we implicate Drosophila Activating transcription factor 3 (Atf3), the single ortholog of human ATF3 and JDP2 bZIP proteins, in abdominal morphogenesis. During the process of the epithelial cell replacement, transcription of the atf3 gene declines. When this downregulation is experimentally prevented, the affected LECs accumulate cell-adhesion proteins and their extrusion and replacement with histoblasts are blocked. The abnormally adhering LECs consequently obstruct the closure of the adult abdominal epithelium. This closure defect can be either mimicked and further enhanced by knockdown of the small GTPase Rho1 or, conversely, alleviated by stimulating ecdysone steroid hormone signaling. Both Rho and ecdysone pathways have been previously identified as effectors of the LEC replacement. To elicit the gain-of-function effect, Atf3 specifically requires its binding partner Jun. Our data thus identify Atf3 as a new functional partner of Drosophila Jun during development.

Interaction between Drosophila bZIP proteins Atf3 and Jun prevents replacement of epithelial cells during metamorphosis.

&Eacute;pith&eacute;lial feuille de diffusion et de la fusion sont &agrave; la base des processus de d&eacute;veloppement importants. Exemples bien caract&eacute;ris&eacute;s de tels &eacute;v&eacute;nements morphog&eacute;n&eacute;tiques &eacute;pith&eacute;liales ont &eacute;t&eacute; fournies par des &eacute;tudes chez la drosophile et incluent la fermeture dorsale embryonnaire, formation du thorax adulte et la cicatrisation. Tous ces processus n&eacute;cessitent les facteurs de transcription base r&eacute;gion-leucine zipper (bZIP) Jun et Fos. Beaucoup moins on conna&icirc;t de la morphogen&egrave;se de l&#39;abdomen de mouche, qui implique le remplacement des cellules &eacute;pidermiques larvaires (ESL) avec les histoblasts adultes qui divisent, migrer et enfin fusionnent pour former l&#39;&eacute;piderme adulte durant la m&eacute;tamorphose. Ici, nous impliquent le facteur de transcription activation de drosophile 3 (Atf3), l&#39;orthologue unique de prot&eacute;ines de bZIP humaines ATF3 et JDP2, dans la morphogen&egrave;se des abdominale. Au cours du processus du remplacement des cellules &eacute;pith&eacute;liales, la transcription du g&egrave;ne atf3 d&eacute;cline. Lorsque cette diminution de l&#39;expression est emp&ecirc;ch&eacute;e exp&eacute;rimentalement, les ESL touch&eacute;s s&#39;accumulent des prot&eacute;ines d&#39;adh&eacute;sion cellulaire et leur extrusion et son remplacement par histoblasts sont bloqu&eacute;s. L&#39;ESL anormalement adh&eacute;rentes en cons&eacute;quence entraver la fermeture de l&#39;&eacute;pith&eacute;lium abdominale adulte. Ce d&eacute;faut de fermeture peut &ecirc;tre imit&eacute; et rehauss&eacute; par la pr&eacute;cipitation de la petite GTPase Rho1 ou, inversement, all&eacute;g&eacute; en stimulant l&#39;ecdysone hormone st&eacute;ro&iuml;de de signalisation. Rho tant l&#39;ecdysone voies ont &eacute;t&eacute; pr&eacute;c&eacute;demment identifi&eacute;s comme effecteurs de la substitution de l&#39;ESL. Pour obtenir l&#39;effet de gain de fonction, Atf3 pr&eacute;voit express&eacute;ment que son partenaire de liaison Jun. Nos donn&eacute;es ainsi identifient Atf3 comme nouveau partenaire fonctionnel de Drosophila Jun au cours du d&eacute;veloppement.

Interaction entre Drosophila bZIP prot&eacute;ines Atf3 et Jun emp&ecirc;che le remplacement des cellules &eacute;pith&eacute;liales durant la m&eacute;tamorphose.

Foglio epiteliale diffusione e fusione sono alla base di importanti processi di sviluppo. Esempi ben caratterizzati di tali eventi morfogenetici epiteliali sono stati forniti da studi in Drosophila e includono chiusura dorsale embrionale, formazione del torace degli adulti e la guarigione delle ferite. Tutti questi processi richiedono i fattori di trascrizione di base regione-leucina zipper (bZIP) Jun e Fos. Molto meno &egrave; conosciuto circa morfogenesi dell&#39;addome vola, che comporta la sostituzione delle cellule epidermiche larvale (LECs) con histoblasts adulti che dividono, migrare e infine si fondono per formare l&#39;epidermide degli adulti durante la metamorfosi. Qui, ci coinvolgono drosofila attivando la fattore di trascrizione 3 (Atf3), il singolo ortholog di ATF3 e JDP2 bZIP proteine umane, nella morfogenesi addominale. Durante il processo di sostituzione delle cellule epiteliali, la trascrizione del gene atf3 declina. Quando questa downregulation sperimentalmente &egrave; impedita, il LECs colpite si accumulano proteine di adesione cellulare e loro sostituzione con histoblasts ed estrusione sono bloccati. LECs anormalmente aderente di conseguenza ostacolare la chiusura dell&#39;epitelio addominale adulto. Questo difetto di chiusura pu&ograve; essere imitato e ulteriormente rafforzato da atterramento del piccolo GTPase Rho1 o, al contrario, alleviato da stimolante segnalazione dell&#39;ormone steroideo ecdisone. Vie ecdisone e di Rho sono stati identificati in precedenza come effettori della sostituzione di LEC. Per suscitare l&#39;effetto di guadagno di funzione, Atf3 richiede specificamente relativo socio associazione giu. I nostri dati cos&igrave; identificano Atf3 come nuovo partner funzionale della drosofila giu durante lo sviluppo.

Interazione tra drosofila bZIP proteine Atf3 e giu impedisce la sostituzione delle cellule epiteliali durante la metamorfosi.

Hoja epitelial separarse y fusi&oacute;n subyacen en importantes procesos de desarrollo. Ejemplos bien caracterizados de acontecimientos morfogen&eacute;ticos epiteliales han sido proporcionados por estudios en Drosophila, incluyen cierre dorsal embrionaria, formaci&oacute;n del t&oacute;rax para adultos y la cicatrizaci&oacute;n de heridas. Todos estos procesos requieren los factores de la transcripci&oacute;n de la cremallera (bZIP) b&aacute;sica de la regi&oacute;n-leucina Jun y Fos. Mucho menos se sabe sobre la morfog&eacute;nesis de la mosca del abdomen, que implica el reemplazo de las c&eacute;lulas epid&eacute;rmicas larvas (LECs) con histoblasts adultos que dividen, migrar y finalmente se fusionan para formar la epidermis adultos durante la metamorfosis. Aqu&iacute;, nos implican Drosophila activando el factor de transcripci&oacute;n 3 (Atf3), el &uacute;nico ort&oacute;logo de prote&iacute;nas bZIP humanas de ATF3 y JDP2, en la morfog&eacute;nesis abdominal. Durante el proceso de la sustituci&oacute;n de c&eacute;lulas epiteliales, la transcripci&oacute;n del gen atf3 declina. Cuando experimentalmente se previene esta regulaci&oacute;n a la baja, los LECs afectados acumulan prote&iacute;nas de adhesi&oacute;n celular y su extrusi&oacute;n y reemplazo con histoblasts est&aacute;n bloqueados. Los LECs anormalmente adherentes en consecuencia obstruyan el cierre del epitelio abdominal para adultos. Este defecto de cierre puede m&iacute;mico y reforzada por la ca&iacute;da de la peque&ntilde;a GTPase Rho1 o, por el contrario, aliviada por la se&ntilde;alizaci&oacute;n de hormona esteroide Ecdisona estimulante. Rho y Ecdisona v&iacute;as han sido previamente identificadas como efectores de la sustituci&oacute;n de la LEC. Para obtener el efecto de ganancia de funci&oacute;n, Atf3 requiere espec&iacute;ficamente su socio enlace Jun. Nuestros datos as&iacute; identifican Atf3 como un nuevo socio funcional de Drosophila Jun durante el desarrollo.

Interacci&oacute;n entre Drosophila bZIP prote&iacute;nas Atf3 y Jun evita el reemplazo de las c&eacute;lulas epiteliales durante la metamorfosis.

FGF signaling is a central regulator of branching morphogenesis processes, such as angiogenesis or the development of branched organs including lung, kidney, and mammary gland. The formation of the air sac during the development of the Drosophila tracheal system is a powerful genetic model to investigate how FGF signaling patterns such emerging structures. This article describes the characterization of the Drosophila matrix metalloprotease Mmp2 as an extracellular inhibitor of FGF morphogenetic function. Mmp2 expression in the developing air sac is controlled by the Drosophila FGF homolog Branchless and then participates in a negative feedback and lateral inhibition mechanism that defines the precise pattern of FGF signaling. The signaling function for MMPs described here may not be limited to branching morphogenesis processes.

Spatial restriction of FGF signaling by a matrix metalloprotease controls branching morphogenesis.

Signalisation de FGF est un r&eacute;gulateur central de ramification morphog&eacute;n&egrave;se, comme l&#39;angiogen&egrave;se ou le d&eacute;veloppement des organes ramifi&eacute;s, y compris les poumons, les reins et les glandes mammaires. La formation du sac a&eacute;rien au cours du d&eacute;veloppement du syst&egrave;me trach&eacute;al drosophile est un puissant mod&egrave;le g&eacute;n&eacute;tique pour &eacute;tudier comment FGF signalisation patterns de telles structures &eacute;mergentes. Cet article d&eacute;crit la caract&eacute;risation de la m&eacute;talloprot&eacute;ase matricielle de Drosophila Mmp2 comme un inhibiteur extracellulaire de fonction morphog&eacute;n&eacute;tique FGF. Expression Mmp2 dans le sac d&#39;air en d&eacute;veloppement est contr&ocirc;l&eacute;e par l&#39;homologue de Drosophila FGF Branchless et participe ensuite &agrave; une r&eacute;troaction n&eacute;gative et le m&eacute;canisme d&#39;inhibition lat&eacute;rale qui d&eacute;finit le mod&egrave;le pr&eacute;cis de la FGF signalisation. La fonction de signalisation pour MPM d&eacute;crit ici n&#39;est peut-&ecirc;tre pas limit&eacute;e &agrave; la ramification des processus de morphogen&egrave;se.

Restriction spatiale de la FGF signalisation par une m&eacute;talloprot&eacute;ase matricielle contr&ocirc;le ramification de la morphogen&egrave;se.

FGF di segnalazione &egrave; un regolatore centrale dei processi di morfogenesi ramificazione, quali l&#39;angiogenesi o lo sviluppo di organi ramificati tra cui polmone, rene e ghiandola mammaria. La formazione del sac di aria durante lo sviluppo del sistema trachea della drosofila &egrave; un potente modello genetico per indagare come segnalazione FGF modelli tali strutture emergenti. Questo articolo descrive la caratterizzazione della drosofila matrix metalloprotease Mmp2 come inibitore extracellulare della funzione morfogenetica FGF. Mmp2 espressione in sviluppo sac di aria &egrave; controllata dall&#39;omologo della drosofila FGF Branchless e poi partecipa a un feedback negativo e il meccanismo di inibizione laterale che definisce il modello preciso di FGF segnalando. La funzione di segnalazione per MMP qui descritto non pu&ograve; essere limitata ai processi di morfogenesi di ramificazione.

Restrizione spaziale di FGF segnalando da una matrice metalloprotease controlla la morfogenesi di ramificazione.

Se&ntilde;alizaci&oacute;n de FGF es un regulador central de ramificaci&oacute;n de los procesos de morfog&eacute;nesis, como angiog&eacute;nesis o el desarrollo de &oacute;rganos ramificados como pulm&oacute;n, ri&ntilde;&oacute;n y gl&aacute;ndula mamaria. La formaci&oacute;n del saco de aire durante el desarrollo del sistema traqueal de Drosophila es un poderoso modelo gen&eacute;tico para investigar c&oacute;mo FGF Se&ntilde;alizaci&oacute;n patrones tales estructuras emergentes. Este art&iacute;culo describe la caracterizaci&oacute;n de la metaloproteinasa de la matriz de Drosophila Mmp2 como inhibidor de la funci&oacute;n morfogen&eacute;tica FGF extracelular. Expresi&oacute;n de Mmp2 en el saco de aire en desarrollo es controlado por el hom&oacute;logo de Drosophila FGF Branchless y participa en un mecanismo de inhibici&oacute;n lateral que define el patr&oacute;n exacto de FGF Se&ntilde;alizaci&oacute;n y retroalimentaci&oacute;n negativa. La funci&oacute;n de se&ntilde;alizaci&oacute;n para MMPs descrito aqu&iacute; no puede limitarse a procesos de morfog&eacute;nesis de ramificaci&oacute;n.

Restricci&oacute;n espacial de FGF Se&ntilde;alizaci&oacute;n por una metaloproteinasa de matriz controla la ramificaci&oacute;n de la morfog&eacute;nesis.

Integration of metabolic and immune responses during animal development ensures energy balance, permitting both growth and defense. Disturbed homeostasis causes organ failure, growth retardation, and metabolic disorders. Here, we show that the Drosophila melanogaster activating transcription factor 3 (Atf3) safeguards metabolic and immune system homeostasis. Loss of Atf3 results in chronic inflammation and starvation responses mounted primarily by the larval gut epithelium, while the fat body suffers lipid overload, causing energy imbalance and death. Hyperactive proinflammatory and stress signaling through NF-κB/Relish, Jun N-terminal kinase, and FOXO in atf3 mutants deregulates genes important for immune defense, digestion, and lipid metabolism. Reducing the dose of either FOXO or Relish normalizes both lipid metabolism and gene expression in atf3 mutants. The function of Atf3 is conserved, as human ATF3 averts some of the Drosophila mutant phenotypes, improving their survival. The single Drosophila Atf3 may incorporate the diversified roles of two related mammalian proteins.

Activating transcription factor 3 regulates immune and metabolic homeostasis.

Cell shape dynamics, motility, and cell proliferation all depend on the actin cytoskeleton. Malignant cancer cells hijack the actin network to grow and migrate to secondary sites. Understanding the function of actin regulators is therefore of major interest. In the present study, we identify the actin cross-linking protein Filamin/Cheerio (Cher) as a mediator of malignancy in genetically defined Drosophila tumors. We show that in invasive tumors, resulting from cooperation of activated Ras with disrupted epithelial cell polarity, Cher is upregulated in a Jun N-terminal kinase (JNK)-dependent manner. Although dispensable in normal epithelium, Cher becomes required in the tumor cells for their growth and invasiveness. When deprived of Cher, these tumor clones lose their full potential to proliferate and breach tissue boundaries. Instead, the Cher-deficient clones remain confined within the limits of their source epithelium, permitting survival of the host animal. Through interaction with the myosin II heavy chain subunit, Cher is likely to strengthen the cortical actomyosin network and reinforce mechanical tension within the invasive tumors. Accordingly, Cher is required for aberrant expression of genes downstream of the Hippo/Yorkie signaling in the tumor tissue. Our study identifies Cher as a new target of JNK signaling that links cytoskeleton dynamics to tumor progression.

The actin cross-linker Filamin/Cheerio mediates tumor malignancy downstream of JNK signaling.

The steroid hormone ecdysone coordinates insect growth and development, directing the major postembryonic transition of forms, metamorphosis. The steroid-deficient ecdysoneless1 (ecd1) strain of Drosophila melanogaster has long served to assess the impact of ecdysone on gene regulation, morphogenesis, or reproduction. However, ecd also exerts cell-autonomous effects independently of the hormone, and mammalian Ecd homologs have been implicated in cell cycle regulation and cancer. Why the Drosophila ecd1 mutants lack ecdysone has not been resolved. Here, we show that in Drosophila cells, Ecd directly interacts with core components of the U5 snRNP spliceosomal complex, including the conserved Prp8 protein. In accord with a function in pre-mRNA splicing, Ecd and Prp8 are cell-autonomously required for survival of proliferating cells within the larval imaginal discs. In the steroidogenic prothoracic gland, loss of Ecd or Prp8 prevents splicing of a large intron from CYP307A2/spookier (spok) pre-mRNA, thus eliminating this essential ecdysone-biosynthetic enzyme and blocking the entry to metamorphosis. Human Ecd (hEcd) can substitute for its missing fly ortholog. When expressed in the Ecd-deficient prothoracic gland, hEcd re-establishes spok pre-mRNA splicing and protein expression, restoring ecdysone synthesis and normal development. Our work identifies Ecd as a novel pre-mRNA splicing factor whose function has been conserved in its human counterpart. Whether the role of mammalian Ecd in cancer involves pre-mRNA splicing remains to be discovered.

Unexpected role of the steroid-deficiency protein ecdysoneless in pre-mRNA splicing.

Juvenile hormones (JHs) play a major role in controlling development and reproduction in insects and other arthropods. Synthetic JH-mimicking compounds such as methoprene are employed as potent insecticides against significant agricultural, household and disease vector pests. However, a receptor mediating effects of JH and its insecticidal mimics has long been the subject of controversy. The bHLH-PAS protein Methoprene-tolerant (Met), along with its Drosophila melanogaster paralog germ cell-expressed (Gce), has emerged as a prime JH receptor candidate, but critical evidence that this protein must bind JH to fulfill its role in normal insect development has been missing. Here, we show that Gce binds a native D. melanogaster JH, its precursor methyl farnesoate, and some synthetic JH mimics. Conditional on this ligand binding, Gce mediates JH-dependent gene expression and the hormone's vital role during development of the fly. Any one of three different single amino acid mutations in the ligand-binding pocket that prevent binding of JH to the protein block these functions. Only transgenic Gce capable of binding JH can restore sensitivity to JH mimics in D. melanogaster Met-null mutants and rescue viability in flies lacking both Gce and Met that would otherwise die at pupation. Similarly, the absence of Gce and Met can be compensated by expression of wild-type but not mutated transgenic D. melanogaster Met protein. This genetic evidence definitively establishes Gce/Met in a JH receptor role, thus resolving a long-standing question in arthropod biology.

Genetic Evidence for Function of the bHLH-PAS Protein Gce/Met As a Juvenile Hormone Receptor.

Cancer initiation and maintenance of the transformed cell state depend on altered cellular signaling and aberrant activities of transcription factors (TFs) that drive pathological gene expression in response to cooperating genetic lesions. Deciphering the roles of interacting TFs is therefore central to understanding carcinogenesis and for designing cancer therapies. Here, we use an unbiased genomic approach to define a TF network that triggers an abnormal gene expression program promoting malignancy of clonal tumors, generated in Drosophila imaginal disc epithelium by gain of oncogenic Ras (Ras(V12)) and loss of the tumor suppressor Scribble (scrib(1)). We show that malignant transformation of the ras(V12)scrib(1) tumors requires TFs of distinct families, namely the bZIP protein Fos, the ETS-domain factor Ets21c and the nuclear receptor Ftz-F1, all acting downstream of Jun-N-terminal kinase (JNK). Depleting any of the three TFs improves viability of tumor-bearing larvae, and this positive effect can be enhanced further by their combined removal. Although both Fos and Ftz-F1 synergistically contribute to ras(V12)scrib(1) tumor invasiveness, only Fos is required for JNK-induced differentiation defects and Matrix metalloprotease (MMP1) upregulation. In contrast, the Fos-dimerizing partner Jun is dispensable for JNK to exert its effects in ras(V12)scrib(1) tumors. Interestingly, Ets21c and Ftz-F1 are transcriptionally induced in these tumors in a JNK- and Fos-dependent manner, thereby demonstrating a hierarchy within the tripartite TF network, with Fos acting as the most upstream JNK effector. Of the three TFs, only Ets21c can efficiently substitute for loss of polarity and cooperate with Ras(V12) in inducing malignant clones that, like ras(V12)scrib(1) tumors, invade other tissues and overexpress MMP1 and the Drosophila insulin-like peptide 8 (Dilp8). While ras(V12)ets21c tumors require JNK for invasiveness, the JNK activity is dispensable for their growth. In conclusion, our study delineates both unique and overlapping functions of distinct TFs that cooperatively promote aberrant expression of target genes, leading to malignant tumor phenotypes.

Mirka Uhlirova

Institute for Genetics,
Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD),
Institute for Genetics, Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD)

University of Cologne

The Drosophila Imaginal Disc Tumor Model: Visualization and Quantification of Gene Expression and Tumor Invasiveness Using Genetic Mosaics