Phagocytosis Assay for Apoptotic Cells in <em>Drosophila</em> Embryos

The molecular mechanisms underlying the phagocytosis of apoptotic cells need to be elucidated in more detail because of its role in immune and ...

Kanazawa University

Lessons from Japanese physicians' education and UK budget increase.

To identify molecules that play roles in the clearance of apoptotic cells by Drosophila phagocytes, we examined a series of monoclonal antibodies raised against larval hemocytes for effects on phagocytosis in vitro. One antibody that inhibited phagocytosis recognized terribly reduced optic lobes (Trol), a core protein of the perlecan-type proteoglycan, and the level of phagocytosis in embryos of a Trol-lacking fly line was lower than in a control line. The treatment of a hemocyte cell line with a recombinant Trol protein containing the amino acid sequence RGD augmented the phosphorylation of focal adhesion kinase, a hallmark of integrin activation. A loss of integrin βν, one of the two β subunits of Drosophila integrin, brought about a reduction in the level of apoptotic cell clearance in embryos. The presence of integrin βν at the surface of embryonic hemocytes was confirmed, and forced expression of integrin βν in hemocytes of an integrin βν-lacking fly line recovered the defective phenotype of phagocytosis. Finally, the level of phagocytosis in a fly line that lacks both integrin βν and Draper, another receptor required for the phagocytosis of apoptotic cells, was lower than that in a fly line lacking either protein. We suggest that integrin βν serves as a phagocytosis receptor responsible for the clearance of apoptotic cells in Drosophila, independent of Draper.

Integrin βν-mediated phagocytosis of apoptotic cells in Drosophila embryos.

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The membrane phospholipid phosphatidylserine is exposed on the cell surface during apoptosis and acts as an eat-me signal in the phagocytosis of apoptotic cells in mammals and nematodes. However, whether this is also true in insects was unclear. When milk fat globule-EGF-factor 8, a phosphatidylserine-binding protein of mammals, was ectopically expressed in Drosophila, the level of phagocytosis was reduced whereas this was not the case for the same protein lacking a domain responsible for the binding to phosphatidylserine. We found that the extracellular region of Draper, an engulfment receptor of Drosophila, binds to phosphatidylserine in an enzyme-linked immunosorbent assay-like solid-phase assay as well as in an assay for surface plasmon resonance. A portion of Draper containing domains EMI and NIM located close to the N-terminus was required for the binding to phosphatidylserine, and a Draper protein lacking this region was not active in Drosophila. Finally, the level of tyrosine-phosphorylated Draper, indicative of the activation of Draper, in a hemocyte-derived cell line was increased after treatment with phosphatidylserine-containing liposome. These results indicated that phosphatidylserine serves as an eat-me signal in the phagocytic removal of apoptotic cells in Drosophila, and that Draper is a phosphatidylserine-binding receptor for phagocytosis.

Phosphatidylserine recognition and induction of apoptotic cell clearance by Drosophila engulfment receptor Draper.

La membrane phospholipidique phosphatidyls&eacute;rine affleure &agrave; la surface des cellules pendant l&#39;apoptosis et agit comme un eat-me signal dans la phagocytose des cellules apoptotiques chez les mammif&egrave;res et les n&eacute;matodes. Toutefois, si cela est aussi vrai chez les insectes n&#39;&eacute;tait pas clair. Quand milk fat globule-EGF-facteur 8, une prot&eacute;ine de liaison de phosphatidyls&eacute;rine de mammif&egrave;res, exprimait de fa&ccedil;on ectopique chez la drosophile, le niveau de phagocytose est r&eacute;duit alors que ce n&#39;&eacute;tait pas le cas pour la m&ecirc;me prot&eacute;ine manque un domaine responsable de la liaison &agrave; la phosphatidyls&eacute;rine. Nous avons constat&eacute; que la r&eacute;gion extracellulaire de Draper, un r&eacute;cepteur d&#39;une dur&eacute;e de drosophile, se lie &agrave; la phosphatidyls&eacute;rine lors d&#39;un test immuno-enzymatique de le phase solide comme dosage aussi bien comme un test de r&eacute;sonance plasmonique de surface. Une partie de Draper contenant des domaines EMI et situ&eacute; &agrave; proximit&eacute; de l&#39;extr&eacute;mit&eacute; N-terminale NIM &eacute;tait requise pour la liaison de phosphatidyls&eacute;rine, et une prot&eacute;ine de Draper, d&eacute;pourvu de cette r&eacute;gion n&#39;&eacute;tait pas active chez la drosophile. Enfin, le niveau de tyrosine phosphoryl&eacute;s Draper, l&#39;indicatif de l&#39;activation de Draper, dans une lign&eacute;e de cellules d&eacute;riv&eacute;es de h&eacute;mocytes augmentait apr&egrave;s le traitement avec les liposomes contenant de phosphatidyls&eacute;rine. Ces r&eacute;sultats indiquent que la phosphatidyls&eacute;rine sert un eat-me signaler le retrait phagocytaire des cellules en apoptose chez la drosophile et que Draper est un r&eacute;cepteur de phosphatidyls&eacute;rine contraignant pour la phagocytose.

Reconnaissance de la phosphatidyls&eacute;rine et l&#39;induction de l&#39;apoptose cellulaire d&eacute;gagement par r&eacute;cepteur d&#39;une dur&eacute;e de Drosophila Draper.

Die Membrane Phospholipid Phosphatidylserin auf der Zelloberfl&auml;che ausgesetzt ist, w&auml;hrend der Apoptose und fungiert als eine Eat-mich Signal in der Phagozytose der Apoptotic Zellen in S&auml;ugetieren und Nematoden. Jedoch, ob dies auch bei Insekten gilt, war unklar. Wenn Milch Fett kugelbl&uuml;te-EGF-Faktor 8, ein Phosphatidylserin-verbindliches Protein der S&auml;ugetiere, wurde in Drosophila ectopically ausgedr&uuml;ckt, das Niveau der Phagozytose wurde reduziert, w&auml;hrend dies nicht der Fall f&uuml;r das gleiche Protein fehlt eine Dom&auml;ne f&uuml;r die Bindung an Phosphatidylserin verantwortlich war. Wir fanden, dass die extrazellul&auml;re Region Draper, ein Engulfment-Rezeptor von Drosophila, an Phosphatidylserin in ein Enzym-Assay-wie Festphasen-Immunoassay auch wie ein Assay f&uuml;r Surface Plasmon Resonance bindet. Ein Teil der Draper, enth&auml;lt Dom&auml;nen EMI und NIM in der N&auml;he des N-Terminus f&uuml;r die Bindung an Phosphatidylserin erforderlich war, und ein Draper-Protein fehlt dieser Region war nicht in Drosophila aktiv. Schlie&szlig;lich das Niveau der Tyrosin-phosphoryliert Draper, ein Indikator der Aktivierung von Draper, in einer Hemocyte abgeleiteten Zelllinie wurde nach der Behandlung mit Phosphatidylserin-haltigen Liposomen erh&ouml;ht. Diese Ergebnisse zeigten, dass Phosphatidylserin dient als eine Eat-mir signalisieren die phagozytische Abbau der Apoptotic Zellen in Drosophila und dass Draper ist ein Phosphatidylserin-Bindung-Rezeptor f&uuml;r Phagozytose.

Phosphatidylserin Anerkennung und Induktion von Apoptotic Zelle Clearance von Drosophila Engulfment Rezeptor Draper.

La fosfatidilserina di fosfolipidi di membrana &egrave; esposto sulla superficie delle cellule durante il apoptosis e agisce come un eat-me segnale nella fagocitosi delle cellule apoptotic in mammiferi e nematodi. Tuttavia, se questo &egrave; vero anche negli insetti era poco chiaro. Quando latte grasso globulo-EGF-fattore 8, una proteina-fosfatidilserina dei mammiferi, ectopically &egrave; stato espresso in Drosophila, il livello di fagocitosi &egrave; stato ridotto, considerando che questo non era il caso per la stessa proteina manca un dominio responsabile per l&#39;associazione di fosfatidilserina. Abbiamo trovato che la regione extracellulare di Draper, un recettore engulfment della drosofila, associa alla fosfatidilserina in un&#39;enzyme-linked immunosorbent assay-come fase solida analisi cos&igrave; come in un saggio per risonanza plasmonica di superficie. Una porzione di Draper contenente domini EMI e NIM si trova vicino al dominio N-terminale &egrave; stato richiesta per l&#39;associazione di fosfatidilserina, e una proteina Draper privo di questa regione non era attiva in drosofila. Infine, il livello di tirosina fosforilato Draper, indicativo dell&#39;attivazione di Draper, in una linea cellulare derivata da hemocyte era aumentata dopo il trattamento con fosfatidilserina contenenti liposomi. Questi risultati hanno indicato che la fosfatidilserina serve come un mangia-me del segnale nella rimozione fagocitaria delle cellule apoptotic in Drosophila, e che Draper &egrave; un recettore di fosfatidilserina-associazione per la fagocitosi.

Riconoscimento della fosfatidilserina e induzione di apoptosi delle cellule liquidazione dal recettore di engulfment drosofila Draper.

&#x7D30;&#x80DE;&#x819C;&#x30EA;&#x30F3;&#x8102;&#x8CEA;&#x306E;&#x30DB;&#x30B9;&#x30D5;&#x30A1;&#x30C1;&#x30B8;&#x30EB;&#x30BB;&#x30EA;&#x30F3;&#x306E;&#x7D30;&#x80DE;&#x8868;&#x9762;&#x306B;&#x306F;&#x30A2;&#x30DD;&#x30C8;&#x30FC;&#x30B7;&#x30B9;&#x4E2D;&#x306B;&#x3055;&#x3089;&#x3055;&#x308C;&#x3066;&#x3044;&#x308B;&#x3057;&#x3001;&#x3068;&#x3057;&#x3066;&#x98DF;&#x3079;&#x308B;&#x884C;&#x70BA;-&#x79C1;&#x306F;&#x54FA;&#x4E73;&#x985E;&#x3068;&#x7DDA;&#x866B;&#x306E;&#x30A2;&#x30DD;&#x30C8;&#x30FC;&#x30B7;&#x30B9;&#x7D30;&#x80DE;&#x306E;&#x8CAA;&#x98DF;&#x80FD;&#x306E;&#x4FE1;&#x53F7;&#x3002;&#x3057;&#x304B;&#x3057;&#x3001;&#x304B;&#x3069;&#x3046;&#x304B;&#x3053;&#x308C;&#x306E;&#x6606;&#x866B;&#x306F;&#x660E;&#x78BA;&#x3060;&#x3063;&#x305F;&#x3067;&#x3059;&#x3002;&#x56E0;&#x5B50;&#x30E2;&#x30EB;&#x30C6;&#x30F3;&#x30B0;&#x30ED;&#x30D3;&#x30E5;&#x30FC;&#x30EB; EGF 8 &#x725B;&#x4E73;&#x306E;&#x8102;&#x80AA;&#x3057;&#x3001;&#x30DB;&#x30B9;&#x30D5;&#x30A1;&#x30C1;&#x30B8;&#x30EB;&#x30BB;&#x30EA;&#x30F3;&#x7D50;&#x5408;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x306E;&#x54FA;&#x4E73;&#x985E;&#x306E;&#x840C;&#x51FA;&#x30B7;&#x30E7;&#x30A6;&#x30B8;&#x30E7;&#x30A6;&#x30D0;&#x30A8;&#x3092;&#x8868;&#x73FE;&#x3057;&#x307E;&#x3057;&#x305F;&#x3001;&#x3053;&#x308C;&#x306E;&#x30DB;&#x30B9;&#x30D5;&#x30A1;&#x30C1;&#x30B8;&#x30EB;&#x30BB;&#x30EA;&#x30F3;&#x306B;&#x30D0;&#x30A4;&#x30F3;&#x30C7;&#x30A3;&#x30F3;&#x30B0;&#x3092;&#x884C;&#x3046;&#x30C9;&#x30E1;&#x30A4;&#x30F3;&#x3092;&#x6B20;&#x3051;&#x3066;&#x3044;&#x308B;&#x540C;&#x3058;&#x86CB;&#x767D;&#x8CEA;&#x306E;&#x305F;&#x3081;&#x306E;&#x30B1;&#x30FC;&#x30B9;&#x3067;&#x306F;&#x306A;&#x304B;&#x3063;&#x305F;&#x306B;&#x5BFE;&#x3057;&#x8CAA;&#x98DF;&#x306E;&#x30EC;&#x30D9;&#x30EB;&#x304C;&#x6E1B;&#x5C11;&#x3057;&#x307E;&#x3057;&#x305F;&#x3002;&#x6211;&#x3005; &#x306F;&#x3001;&#x30FB; &#x30C9;&#x30EC;&#x30A4;&#x30D1;&#x30FC;&#x3001;&#x30B7;&#x30E7;&#x30A6;&#x30B8;&#x30E7;&#x30A6;&#x30D0;&#x30A8;&#x306E;&#x8CAA;&#x98DF;&#x53D7;&#x5BB9;&#x4F53;&#x306E;&#x7D30;&#x80DE;&#x5916;&#x9818;&#x57DF;&#x30DB;&#x30B9;&#x30D5;&#x30A1;&#x30C1;&#x30B8;&#x30EB;&#x30BB;&#x30EA;&#x30F3;&#x306F; elisa &#x306E;&#x8A66;&#x91D1;&#x306E;&#x3088;&#x3046;&#x306A;&#x56FA;&#x76F8;&#x30A2;&#x30C3;&#x30BB;&#x30A4;&#x3067;&#x3082;&#x8868;&#x9762;&#x30D7;&#x30E9;&#x30BA;&#x30E2;&#x30F3;&#x5171;&#x9CF4;&#x306E;&#x305F;&#x3081;&#x306E;&#x8A66;&#x91D1;&#x306E;&#x3088;&#x3046;&#x306B;&#x30D0;&#x30A4;&#x30F3;&#x30C9;&#x3055;&#x308C;&#x3066;&#x3044;&#x308B;&#x3053;&#x3068;&#x3092;&#x767A;&#x898B;&#x3057;&#x307E;&#x3057;&#x305F;&#x3002;EMI &#x3068; N &#x672B;&#x7AEF;&#x8FD1;&#x304F;&#x306B;&#x3042;&#x308B; NIM &#x306E;&#x30C9;&#x30E1;&#x30A4;&#x30F3;&#x3092;&#x542B;&#x3080;&#x30C9;&#x30EC;&#x30FC;&#x30D1;&#x30FC;&#x306E;&#x90E8;&#x5206;&#x304C;&#x30DB;&#x30B9;&#x30D5;&#x30A1;&#x30C1;&#x30B8;&#x30EB;&#x30BB;&#x30EA;&#x30F3;&#x3078;&#x306E;&#x30D0;&#x30A4;&#x30F3;&#x30C9;&#x306B;&#x5FC5;&#x8981;&#x306A;&#x3055;&#x308C;&#x3001;&#x3053;&#x306E;&#x5730;&#x57DF;&#x304C;&#x6B20;&#x3051;&#x3066;&#x30C9;&#x30EC;&#x30FC;&#x30D1;&#x30FC; &#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x304C;&#x30B7;&#x30E7;&#x30A6;&#x30B8;&#x30E7;&#x30A6;&#x30D0;&#x30A8;&#x306E;&#x30A2;&#x30AF;&#x30C6;&#x30A3;&#x30D6;&#x3067;&#x3057;&#x305F;&#x3002;&#x6700;&#x5F8C;&#x306B;&#x3001;&#x30C9;&#x30EC;&#x30A4;&#x30D1;&#x30FC;&#x3044;&#x30C1;&#x30ED;&#x30B7;&#x30F3;&#x30EA;&#x30F3;&#x9178;&#x5316;&#x306E;&#x30EC;&#x30D9;&#x30EB;&#x3001;&#x8840;&#x7403;&#x7531;&#x6765;&#x7D30;&#x80DE;&#x306E;&#x6D3B;&#x6027;&#x5316;&#x30FB; &#x30C9;&#x30EC;&#x30A4;&#x30D1;&#x30FC;&#x3001;&#x793A;&#x3059;&#x30DB;&#x30B9;&#x30D5;&#x30A1;&#x30C1;&#x30B8;&#x30EB;&#x30BB;&#x30EA;&#x30F3;&#x542B;&#x6709;&#x30EA;&#x30DD;&#x30BD;&#x30FC;&#x30E0;&#x306B;&#x3088;&#x308B;&#x6CBB;&#x7642;&#x5F8C;&#x306B;&#x5897;&#x52A0;&#x3057;&#x307E;&#x3057;&#x305F;&#x3002;&#x305D;&#x306E;&#x30DB;&#x30B9;&#x30D5;&#x30A1;&#x30C1;&#x30B8;&#x30EB;&#x30BB;&#x30EA;&#x30F3;&#x6A5F;&#x80FD;&#x3068;&#x3057;&#x3066;&#x98DF;&#x3079;&#x308B;&#x3053;&#x3068;&#x304C;&#x793A;&#x5506;-&#x305D;&#x306E;&#x30C9;&#x30EC;&#x30FC;&#x30D1;&#x30FC;&#x8CAA;&#x98DF;&#x306E;&#x30DB;&#x30B9;&#x30D5;&#x30A1;&#x30C1;&#x30B8;&#x30EB;&#x30BB;&#x30EA;&#x30F3;&#x7D50;&#x5408;&#x53D7;&#x5BB9;&#x4F53;&#x306E;&#x30B7;&#x30B0;&#x30CA;&#x30EB;&#x306E;&#x30B7;&#x30E7;&#x30A6;&#x30B8;&#x30E7;&#x30A6;&#x30D0;&#x30A8;&#x306B;&#x304A;&#x3051;&#x308B;&#x30A2;&#x30DD;&#x30C8;&#x30FC;&#x30B7;&#x30B9;&#x7D30;&#x80DE;&#x306E;&#x8CAA;&#x98DF;&#x9664;&#x53BB;&#x3067;&#x3042;&#x308A;&#x3002;

&#x30DB;&#x30B9;&#x30D5;&#x30A1;&#x30C1;&#x30B8;&#x30EB;&#x30BB;&#x30EA;&#x30F3;&#x8A8D;&#x8B58;&#x3068;&#x8A98;&#x5C0E;&#x30A2;&#x30DD;&#x30C8;&#x30FC;&#x30B7;&#x30B9;&#x7D30;&#x80DE;&#x9664;&#x53BB;&#x306B;&#x3088;&#x308B;&#x30B7;&#x30E7;&#x30A6;&#x30B8;&#x30E7;&#x30A6;&#x30D0;&#x30A8;&#x8CAA;&#x98DF;&#x53D7;&#x5BB9;&#x4F53; Draper &#x306E;&#x3002;

&#xB9C9; &#xC778;&#xC9C0;&#xC9C8; phosphatidylserine apoptosis &#xB3D9;&#xC548; &#xC138;&#xD3EC; &#xD45C;&#xBA74;&#xC5D0; &#xB178;&#xCD9C; &#xB418; &#xACE0; &#xC2DD;&#xC0AC; &#xC5ED;&#xD560;-&#xD3EC;&#xC720;&#xB958; &#xBC0F; nematodes apoptotic &#xC138;&#xD3EC;&#xC758; &#xC2DD; &#xADE0; &#xC791;&#xC6A9;&#xC5D0; &#xC2E0;&#xD638; &#xB098;. &#xC5EC;&#xBD80; &#xC774;&#xAC83;&#xC774; &#xC0AC;&#xC2E4; &#xACE4;&#xCDA9;&#xC5D0;&#xB3C4; &#xD558;&#xB294; &#xAC83;&#xC740; &#xBD84;&#xBA85; &#xD569;&#xB2C8;&#xB2E4;. &#xC6B0;&#xC720; &#xC9C0;&#xBC29; globule EGF &#xD329;&#xD130; 8, &#xD3EC;&#xC720;&#xB958;, phosphatidylserine &#xBC14;&#xC778;&#xB529; &#xB2E8;&#xBC31;&#xC9C8; &#xCD08;&#xD30C;&#xB9AC;&#xC5D0; ectopically &#xD45C;&#xD604; &#xD588;&#xB2E4;, &#xC774;&#xAC83;&#xC774; &#xB3C4;&#xBA54;&#xC778; phosphatidylserine&#xC5D0; &#xBC14;&#xC778;&#xB529;&#xC744; &#xC218;&#xD589; &#xBD80;&#xC871; &#xAC19;&#xC740; &#xB2E8;&#xBC31;&#xC9C8;&#xC758; &#xACBD;&#xC6B0; &#xBC18;&#xBA74;&#xC5D0;, &#xC2DD; &#xADE0; &#xC791;&#xC6A9;&#xC758; &#xC218;&#xC900;&#xC744; &#xAC10;&#xC18C; &#xD588;&#xB2E4;. &#xC6B0;&#xB9AC;&#xB294; &#xB4DC;&#xB808;&#xC774;&#xD37C;, &#xCD08;&#xD30C;&#xB9AC;&#xC758; engulfment &#xC218;&#xC6A9; &#xCCB4;&#xC758; &#xC138;&#xD3EC; &#xC678; &#xC601;&#xC5ED; &#xBD84;&#xC11D; &#xD6A8;&#xC18C; &#xC5F0;&#xACB0; &#xB41C; immunosorbent &#xBD84;&#xC11D; &#xACB0;&#xACFC; &#xAC19;&#xC740; &#xACE0;&#xCCB4; &#xC0C1; &#xACB0;&#xACFC;&#xC5D0; &#xD45C;&#xBA74; &#xD50C;&#xB77C;&#xC2A4;&#xBAAC; &#xACF5;&#xBA85;&#xC5D0; &#xB300; &#xD55C; &#xBD84;&#xC11D; &#xACB0;&#xACFC; &#xAC19;&#xC774; phosphatidylserine &#xBC14;&#xC778;&#xB529;&#xB429;&#xB2C8;&#xB2E4; &#xBC1C;&#xACAC;. &#xB4DC;&#xB808;&#xC774;&#xD37C; EMI-&#xB2D8; N &#xB9D0;&#xB2E8; &#xADFC;&#xCC98;&#xC5D0; &#xC704;&#xCE58;&#xD55C; &#xB3C4;&#xBA54;&#xC778;&#xC5D0; &#xD3EC;&#xD568; &#xB41C; &#xC77C;&#xBD80; phosphatidylserine, &#xBC14;&#xC778;&#xB529;&#xD560; &#xD544;&#xC694; &#xC774;&#xC5C8;&#xACE0; &#xB4DC;&#xB808;&#xC774;&#xD37C; &#xB2E8;&#xBC31;&#xC9C8;&#xC774;&#xC774; &#xC9C0;&#xC5ED;&#xC758; &#xBD80;&#xC871; &#xD55C; &#xCD08;&#xD30C;&#xB9AC;&#xC5D0;&#xC11C; &#xD65C;&#xB3D9;&#xC801; &#xC774;&#xC5C8;&#xB2E4;. &#xB9C8;&#xC9C0;&#xB9C9;&#xC73C;&#xB85C;,&#xC758; &#xC218;&#xC900; &#xD2F0;&#xB85C;&#xC2E0; phosphorylated &#xB4DC;&#xB808;&#xC774;&#xD37C;, hemocyte&#xC5D0;&#xC11C; &#xD30C;&#xC0DD; &#xB41C; &#xC140; &#xB77C;&#xC778;&#xC5D0; &#xB4DC;&#xB808;&#xC774;&#xD37C;, &#xD65C;&#xC131;&#xD654;&#xC758; &#xC9C0;&#xD45C;&#xB85C; phosphatidylserine &#xB4E4;&#xC5B4; &#xC788;&#xB294; liposome &#xCE58;&#xB8CC; &#xD6C4; &#xC99D;&#xAC00; &#xD588;&#xB2E4;. &#xC774;&#xB7EC;&#xD55C; &#xACB0;&#xACFC; &#xBA39;&#xACE0; &#xADF8; phosphatidylserine &#xC5ED;&#xD560; &#xD45C;&#xC2DC;-&#xB0A0; apoptotic &#xC138;&#xD3EC; &#xCD08;&#xD30C;&#xB9AC;, phagocytic &#xC81C;&#xAC70;&#xC5D0;&#xC11C; &#xC2E0;&#xD638; &#xC774;&#xBA70; &#xADF8; &#xB4DC;&#xB808;&#xC774;&#xD37C; phosphatidylserine &#xBC14;&#xC778;&#xB529; &#xC218;&#xC6A9; &#xCCB4;&#xC5D0; &#xB300; &#xD55C; &#xC2DD; &#xADE0; &#xC791;&#xC6A9;.

Phosphatidylserine &#xC778;&#xC2DD;&#xACFC; apoptotic &#xC138;&#xD3EC; &#xD074;&#xB9AC;&#xC5B4;&#xB7F0;&#xC2A4; Drosophila engulfment &#xC218;&#xC6A9; &#xCCB4; &#xB4DC;&#xB808;&#xC774;&#xD37C;&#xC5D0; &#xC758;&#xD574; &#xC720;&#xB3C4;.

La fosfatidilserina de fosfol&iacute;pidos de la membrana est&aacute; expuesta en la superficie celular durante la apoptosis y act&uacute;a como una coma-me se&ntilde;al en la fagocitosis de c&eacute;lulas apopt&oacute;ticas en mam&iacute;feros y nematodos. Sin embargo, si esto sucede tambi&eacute;n en insectos era confusa. Cuando el gl&oacute;bulo-EGF-factor 8 de grasa de leche, una prote&iacute;na de uni&oacute;n a fosfatidilserina de mam&iacute;feros, fue expresada ect&oacute;picamente en Drosophila, se redujo el nivel de la fagocitosis mientras que esto no era el caso de la misma prote&iacute;na que carece de un dominio responsable de la Uni&oacute;n a la fosfatidilserina. Encontramos que la regi&oacute;n extracelular de Draper, un receptor de inmersi&oacute;n de Drosophila, se une a la fosfatidilserina en un enzyme-linked immunosorbent ensayo-como fase s&oacute;lida ensayo as&iacute; como un ensayo para resonancia de plasmones de superficie. Una porci&oacute;n de Draper que contiene dominios EMI y situado cerca de la N-terminal NIM fue requerida para el enlace a la fosfatidilserina y una prote&iacute;na de Draper que carecen de esta regi&oacute;n no era activa en Drosophila. Por &uacute;ltimo, el nivel de tirosina-fosforilada Draper, indicativo de la activaci&oacute;n de Draper, en una l&iacute;nea de c&eacute;lulas hemocyte aument&oacute; despu&eacute;s del tratamiento con liposomas que contienen fosfatidilserina. Estos resultados indicaron que fosfatidilserina sirve como una coma-me de la se&ntilde;al en el retiro fagocitario de c&eacute;lulas apopt&oacute;ticas en Drosophila, y eso Draper es un receptor de fosfatidilserina vinculante para la fagocitosis.

Reconocimiento de la fosfatidilserina y la inducci&oacute;n de apoptosis celular separaci&oacute;n por receptores de inmersi&oacute;n de Drosophila Draper.

Integrins exert a variety of cellular functions as heterodimers of two transmembrane subunits named α and β. Integrin βν, a β-subunit of Drosophila integrin, is involved in the phagocytosis of apoptotic cells and bacteria. Here, we searched for an α-subunit that forms a complex and cooperates with βν. Examinations of RNAi-treated animals suggested that αPS3, but not any of four other α-subunits, is required for the effective phagocytosis of apoptotic cells in Drosophila embryos. The mutation of αPS3-encoding scb, deficiency, insertion of P-element, or alteration of nucleotide sequences, brought about a reduction in the level of phagocytosis. The defect in phagocytosis by deficiency was reverted by the forced expression of scb. Furthermore, flies in which the expression of both αPS3 and βν was inhibited by RNAi showed a level of phagocytosis almost equal to that observed in flies with RNAi for either subunit alone. A loss of αPS3 also decreased the activity of larval hemocytes in the phagocytosis of Staphylococcus aureus. Finally, a co-immunoprecipitation analysis using a Drosophila cell line treated with a chemical cross-linker suggested a physical association between αPS3 and βν. These results collectively indicated that integrin αPS3/βν serves as a receptor in the phagocytosis of apoptotic cells and bacteria by Drosophila phagocytes.

Integrin αPS3/βν-mediated phagocytosis of apoptotic cells and bacteria in Drosophila.

Autobiographical memories can be recalled from either a field (first-person) or an observer (third-person) perspective. Previous studies have reported that field-to-observer perspective change reduced the emotional intensity of recalled events. In the present study, we examined whether this effect has a long duration by employing follow-up measurements. The participants were asked to recall the same events repeatedly across three sessions (S1, S2, and S3): S2 was conducted about three days after S1, and S3 was conducted about four weeks after S2. The results showed a reduction in the emotional intensity of the recalled events when the perspective was changed from field to observer at S2. More importantly, this reduction in emotional intensity persisted until S3. These effects were not observed under observer-to-field perspective change at S2. These results suggest that observer perspective taking can cause plastic change in the autobiographical memory system.

The long-term effect of perspective change on the emotional intensity of autobiographical memories.

The phagocytic elimination of cells undergoing apoptosis is an evolutionarily conserved innate immune mechanism for eliminating unnecessary cells. Previous studies showed an increase in the level of engulfment receptors in phagocytes after the phagocytosis of apoptotic cells, which leads to the enhancement of their phagocytic activity. However, precise mechanisms underlying this phenomenon require further clarification. We found that the pre-incubation of a Drosophila phagocyte cell line with the fragments of apoptotic cells enhanced the subsequent phagocytosis of apoptotic cells, accompanied by an augmented expression of the engulfment receptors Draper and integrin αPS3. The DNA-binding activity of the transcription repressor Tailless was transiently raised in those phagocytes, depending on two partially overlapping signal-transduction pathways for the induction of phagocytosis as well as the occurrence of engulfment. The RNAi knockdown of tailless in phagocytes abrogated the enhancement of both phagocytosis and engulfment receptor expression. Furthermore, the hemocyte-specific RNAi of tailless reduced apoptotic cell clearance in Drosophila embryos. Taken together, we propose the following mechanism for the activation of Drosophila phagocytes after an encounter with apoptotic cells: two partially overlapping signal-transduction pathways for phagocytosis are initiated; transcription repressor Tailless is activated; expression of engulfment receptors is stimulated; and phagocytic activity is enhanced. This phenomenon most likely ensures the phagocytic elimination of apoptotic cells by stimulated phagocytes and is thus considered as a mechanism to prime phagocytes in innate immunity.

Saori Nonaka

Graduate School of Medical Sciences

Kanazawa University

Phagocytosis Assay for Apoptotic Cells in Drosophila Embryos

Saori Nonaka

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Graduate School of Medical Sciences

Kanazawa University

Phagocytosis Assay for Apoptotic Cells in Drosophila Embryos

Graduate School of Medical Sciences