collection-analysis-arabidopsis-phloem-exudates-using-edta

Collection and Analysis of <em>Arabidopsis</em> Phloem Exudates Using the EDTA-facilitated Method

The plant phloem is essential for the long-distance transport of (photo-) assimilates as well as of signals conveying biotic or abiotic stress. It ...

Michigan State Universtiy

Physiological evidence indicates that flower formation is hormonally controlled. The floral stimulus, or florigen, is formed in the leaves as a response to an inductive photoperiod and translocated through the phloem to the apical meristem. However, because of difficulties in obtaining and analyzing phloem sap and the lack of a bioassay, the chemical nature of this stimulus is one of the major unsolved problems in plant biology. A combination of microbore high-performance liquid chromatography (HPLC) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) was used to compare the contents of the phloem sap from flowering and non-flowering plants. Instead of using one- or two-dimensional gel electrophoresis, microbore HPLC separations allowed us to detect proteins/peptides that were very small and present at very low levels. We detected more than 100 components in the phloem sap of Perilla ocymoides L. and Lupinus albusL. Sequences for 16 peptides in a mass range from 1 to 9 kDa were obtained. Two of these could be identified, 11 showed similarity to known or deduced protein sequences, and three showed no similarity to any known protein or translated gene sequence. Four of these peptides were specific to, modified, or increased in plants that were flowering, indicating their possible role in flower induction. The sequences of these peptides showed similarities to two purine permeases, a protein with similarity to protein kinases, and a protein with no similarities to any known protein.

Comparison of peptides in the phloem sap of flowering and non-flowering Perilla and lupine plants using microbore HPLC followed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

&#x6269;&#x5C55;&#x86CB;&#x767D;&#x662F;&#x4EE5;&#x7279;&#x8272;&#x548C;&#x72EC;&#x7279;&#x7684;&#x65B9;&#x5F0F;&#x8BF1;&#x4F7F; pH &#x4F9D;&#x8D56;&#x578B;&#x5899;&#x6269;&#x5C55;&#x548C;&#x5E94;&#x529B;&#x677E;&#x5F1B;&#x7684;&#x7EC6;&#x80DE;&#x58C1;&#x86CB;&#x767D;&#x8D28;&#x3002;&#x4E24;&#x4E2A;&#x5BB6;&#x5EAD;&#x7684;&#x6269;&#x5C55;&#x86CB;&#x767D;&#x4F17;&#x6240;&#x5468;&#x77E5;&#x7684;&#x540D;&#x4E3A;&#x6269;&#x5C55; &alpha;-&#x548C; &beta;-&#x86CB;&#x767D;&#xFF0C;&#x548C;&#x4ED6;&#x4EEC;&#x7EC4;&#x6210;&#x5927;&#x578B;&#x9176;&#x5BB6;&#x5EAD;&#xFF0C;&#x5176;&#x6210;&#x5458;&#x79C0;&#x5404;&#x79CD;&#x5668;&#x5B98;&#x3001; &#x7EC4;&#x7EC7;&#x548C;&#x7EC6;&#x80DE;&#x7279;&#x5F02;&#x6027;&#x8868;&#x8FBE;&#x6A21;&#x5F0F;&#x3002;&#x5176;&#x4ED6;&#x6269;&#x5C55;&#x86CB;&#x767D;&#x9065;&#x8FDC;&#x5E8F;&#x5217;&#x76F8;&#x4F3C;&#x7684;&#x57FA;&#x56E0;&#x4E5F;&#x6709;&#x4EE3;&#x8868;&#x5728;&#x5E8F;&#x5217;&#x6570;&#x636E;&#x5E93;&#x4E2D;&#xFF0C;&#x4F46;&#x5176;&#x751F;&#x7269;&#x548C;&#x751F;&#x5316;&#x529F;&#x80FD;&#x5C1A;&#x672A;&#x53D1;&#x73B0;&#x3002;&#x4F38;&#x5C55;&#x86CB;&#x767D;&#x4F3C;&#x4E4E;&#x524A;&#x5F31;&#x8461;&#x805A;&#x7CD6;-&#x8461;&#x805A;&#x7CD6;&#x5177;&#x6709;&#x7EA6;&#x675F;&#x529B;&#xFF0C;&#x4F46;&#x5176;&#x8BE6;&#x7EC6;&#x7684;&#x673A;&#x5236;&#x7684;&#x884C;&#x52A8;&#x8FD8;&#x672A;&#x5B8C;&#x5168;&#x786E;&#x7ACB;&#x3002;&#x6269;&#x5C55;&#x86CB;&#x767D;&#x7684;&#x751F;&#x7269;&#x5B66;&#x4F5C;&#x7528;&#x662F;&#x591A;&#x79CD;&#x591A;&#x6837;&#x7684;&#x4F46;&#x53EF;&#x4EE5;&#x6269;&#x5C55;&#x86CB;&#x767D;&#x677E;&#x5F00;&#x7EC6;&#x80DE;&#x58C1;&#xFF0C;&#x4F8B;&#x5982;&#x5728;&#x7EC6;&#x80DE;&#x589E;&#x5927;&#x3001; &#x679C;&#x5B9E;&#x8F6F;&#x5316;&#x3001; &#x82B1;&#x7C89;&#x7BA1;&#x53CA;&#x6839;&#x5934;&#x53D1;&#x751F;&#x957F;&#x548C;&#x8131;&#x843D;&#x7684;&#x884C;&#x52A8;&#x6709;&#x5173;&#x3002;&#x4F38;&#x5C55;&#x86CB;&#x767D;&#x6837;&#x86CB;&#x767D;&#x4E5F;&#x67E5;&#x51FA;&#x7EC6;&#x83CC;&#x548C;&#x771F;&#x83CC;&#xFF0C;&#x4ED6;&#x4EEC;&#x53EF;&#x80FD;&#x4F1A;&#x5728;&#x5176;&#x4E2D;&#x5E2E;&#x52A9;&#x690D;&#x7269;&#x4F53;&#x5185;&#x7684;&#x5FAE;&#x751F;&#x7269;&#x5165;&#x4FB5;&#x3002;

&#x6269;&#x5C55;&#x86CB;&#x767D;&#x98DE;&#x901F;&#x53D1;&#x5C55;&#x7684;&#x4E16;&#x754C;&#x3002;

Expansins are cell wall proteins that induce pH-dependent wall extension and stress relaxation in a characteristic and unique manner. Two families of expansins are known, named alpha- and beta-expansins, and they comprise large multigene families whose members show diverse organ-, tissue- and cell-specific expression patterns. Other genes that bear distant sequence similarity to expansins are also represented in the sequence databases, but their biological and biochemical functions have not yet been uncovered. Expansin appears to weaken glucan-glucan binding, but its detailed mechanism of action is not well established. The biological roles of expansins are diverse, but can be related to the action of expansins to loosen cell walls, for example during cell enlargement, fruit softening, pollen tube and root hair growth, and abscission. Expansin-like proteins have also been identified in bacteria and fungi, where they may aid microbial invasion of the plant body.

The growing world of expansins.

Expansines sont des prot&#xE9;ines de parois cellulaires qui induisent d&#xE9;pendant du pH d&#x26;#39;extension paroi et la relaxation de contrainte d&#x26;#39;une mani&#xE8;re caract&#xE9;ristique et unique. Deux familles de expansines sont connus, le nom d&#x26;#39;alpha-et b&#xEA;ta-expansines, et ils comprennent de grandes familles multig&#xE9;niques dont les membres montrent diverses organe, au tissu et mod&#xE8;les cellulaires sp&#xE9;cifiques d&#x26;#39;expression. D&#x26;#39;autres g&#xE8;nes qui portent une similarit&#xE9; de s&#xE9;quence distant pour expansines sont &#xE9;galement repr&#xE9;sent&#xE9;s dans les bases de donn&#xE9;es de s&#xE9;quences, mais leurs fonctions biologiques et biochimiques n&#x26;#39;ont pas encore &#xE9;t&#xE9; d&#xE9;couvert. Expansine semble affaiblir glucane-liaison au glucane, mais son m&#xE9;canisme d&#x26;#39;action d&#xE9;taill&#xE9; n&#x26;#39;est pas bien &#xE9;tablie. Les r&#xF4;les biologiques des expansines sont diverses, mais peut &#xEA;tre li&#xE9;e &#xE0; l&#x26;#39;action de expansines pour desserrer les parois des cellules, par exemple lors d&#x26;#39;une hypertrophie des cellules, ramollissement des fruits, du tube pollinique et la croissance des poils racinaires, et l&#x26;#39;abscission. Expansine-comme les prot&#xE9;ines ont &#xE9;galement &#xE9;t&#xE9; identifi&#xE9;s chez les bact&#xE9;ries et les champignons, o&#xF9; ils peuvent aider l&#x26;#39;invasion microbienne de corps de la plante.

Le monde en croissance de expansines

Die wachsende Welt der Expansinen.

Expansins sono le proteine della parete cellulare che inducono rilassamento estensione e lo stress di parete pH-dipendente in modo caratteristico e unico. Sono noti due famiglie di expansins, denominata alfa - e beta-expansins, e comprendono due grandi multigene famiglie i cui membri mostrano diverse organo-, modelli di espressione del tessuto e delle cellule specifiche. Altri geni che portano la somiglianza di sequenza lontane a expansins sono anche rappresentati nei database di sequenza, ma le loro funzioni biologiche e biochimiche non sono ancora stati scoperti. Expansin sembra indebolire glucano-glucano vincolante, ma suo dettagliato meccanismo d&#39;azione non &egrave; ben definito. I ruoli biologici di expansins sono diversi, ma possono essere correlati all&#39;azione di expansins per allentare pareti della cellula, per esempio durante l&#39;allargamento delle cellule, frutta rammollimento, tubo pollinico e la crescita dei capelli radice e abscissione. Expansin-come le proteine sono state identificate anche in batteri e funghi, dove si pu&ograve; essere di aiuto invasione microbica del corpo vegetale.

Il mondo crescente di expansins.

&#x30A8;&#x30AF;&#x30B9;&#x30D1;&#x30F3;&#x30B7;&#x30F3;&#x306F;&#x3001;&#x7279;&#x6027;&#x3068;&#x72EC;&#x81EA;&#x306E;&#x65B9;&#x6CD5;&#x3067;pH&#x4F9D;&#x5B58;&#x6027;&#x306E;&#x58C1;&#x306E;&#x62E1;&#x5F35;&#x3068;&#x5FDC;&#x529B;&#x7DE9;&#x548C;&#x3092;&#x8A98;&#x767A;&#x3059;&#x308B;&#x7D30;&#x80DE;&#x58C1;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x3067;&#x3042;&#x308B;&#x3002;&#x30A8;&#x30AF;&#x30B9;&#x30D1;&#x30F3;&#x30B7;&#x30F3;&#x306E;2&#x3064;&#x306E;&#x30D5;&#x30A1;&#x30DF;&#x30EA;&#x30FC;&#x306F;&#x3001;&#x77E5;&#x3089;&#x308C;&#x3066;&#x3044;&#x308B;&#x3B1;-&#x304A;&#x3088;&#x3073;&#x3B2;-&#x30A8;&#x30AF;&#x30B9;&#x30D1;&#x30F3;&#x30B7;&#x30F3;&#x306E;&#x540D;&#x524D;&#x3001;&#x305D;&#x3057;&#x3066;&#x3001;&#x5F7C;&#x3089;&#x306F;&#x305D;&#x306E;&#x30E1;&#x30F3;&#x30D0;&#x30FC;&#x306E;&#x591A;&#x69D8;&#x306A;&#x81D3;&#x5668;&#x3001;&#x7D44;&#x7E54;&#x304A;&#x3088;&#x3073;&#x7D30;&#x80DE;&#x7279;&#x7570;&#x7684;&#x767A;&#x73FE;&#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x3092;&#x793A;&#x3059;&#x5927;&#x898F;&#x6A21;&#x306A;&#x591A;&#x91CD;&#x907A;&#x4F1D;&#x5B50;&#x30D5;&#x30A1;&#x30DF;&#x30EA;&#x30FC;&#x3092;&#x69CB;&#x6210;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x30A8;&#x30AF;&#x30B9;&#x30D1;&#x30F3;&#x30B7;&#x30F3;&#x306E;&#x9060;&#x3044;&#x914D;&#x5217;&#x985E;&#x4F3C;&#x6027;&#x3092;&#x8CA0;&#x62C5;&#x3059;&#x308B;&#x4ED6;&#x306E;&#x907A;&#x4F1D;&#x5B50;&#x306F;&#x3001;&#x914D;&#x5217;&#x30C7;&#x30FC;&#x30BF;&#x30D9;&#x30FC;&#x30B9;&#x3067;&#x8868;&#x3055;&#x308C;&#x308B;&#x304C;&#x3001;&#x305D;&#x306E;&#x751F;&#x7269;&#x5B66;&#x7684;&#x304A;&#x3088;&#x3073;&#x751F;&#x5316;&#x5B66;&#x7684;&#x6A5F;&#x80FD;&#x306F;&#x307E;&#x3060;&#x660E;&#x3089;&#x304B;&#x306B;&#x3055;&#x308C;&#x3066;&#x3044;&#x306A;&#x3044;&#x3002;&#x30A8;&#x30AF;&#x30B9;&#x30D1;&#x30F3;&#x30B7;&#x30F3;&#x306F;&#x3001;&#x30B0;&#x30EB;&#x30AB;&#x30F3;&#x30B0;&#x30EB;&#x30AB;&#x30F3;&#x306E;&#x7D50;&#x5408;&#x3092;&#x5F31;&#x3081;&#x308B;&#x3088;&#x3046;&#x306B;&#x898B;&#x3048;&#x307E;&#x3059;&#x304C;&#x3001;&#x305D;&#x306E;&#x4F5C;&#x7528;&#x6A5F;&#x69CB;&#x306E;&#x8A73;&#x7D30;&#x306F;&#x5341;&#x5206;&#x306B;&#x78BA;&#x7ACB;&#x3055;&#x308C;&#x3066;&#x3044;&#x307E;&#x305B;&#x3093;&#x3002;&#x30A8;&#x30AF;&#x30B9;&#x30D1;&#x30F3;&#x30B7;&#x30F3;&#x306E;&#x751F;&#x7269;&#x5B66;&#x7684;&#x5F79;&#x5272;&#x306F;&#x591A;&#x69D8;&#x3067;&#x3042;&#x308B;&#x304C;&#x3001;&#x7D30;&#x80DE;&#x80A5;&#x5927;&#x3001;&#x679C;&#x5B9F;&#x306E;&#x8EDF;&#x5316;&#x3001;&#x82B1;&#x7C89;&#x7BA1;&#x3068;&#x6839;&#x6BDB;&#x306E;&#x6210;&#x9577;&#x3001;&#x8131;&#x96E2;&#x6642;&#x306B;&#x3001;&#x4F8B;&#x3048;&#x3070;&#x3001;&#x7D30;&#x80DE;&#x58C1;&#x3092;&#x7DE9;&#x3081;&#x308B;&#x305F;&#x3081;&#x306B;&#x30A8;&#x30AF;&#x30B9;&#x30D1;&#x30F3;&#x30B7;&#x30F3;&#x306E;&#x4F5C;&#x7528;&#x306B;&#x95A2;&#x9023;&#x3059;&#x308B;&#x3053;&#x3068;&#x304C;&#x3067;&#x304D;&#x307E;&#x3059;&#x3002;&#x30A8;&#x30AF;&#x30B9;&#x30D1;&#x30F3;&#x30B7;&#x30F3;&#x306E;&#x3088;&#x3046;&#x306A;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x306F;&#x307E;&#x305F;&#x3001;&#x5F7C;&#x3089;&#x306F;&#x690D;&#x7269;&#x4F53;&#x306E;&#x5FAE;&#x751F;&#x7269;&#x306E;&#x4FB5;&#x5165;&#x3092;&#x52A9;&#x3051;&#x308B;&#x304B;&#x3082;&#x3057;&#x308C;&#x306A;&#x3044;&#x3068;&#x3053;&#x308D;&#x3001;&#x7D30;&#x83CC;&#x304A;&#x3088;&#x3073;&#x771F;&#x83CC;&#x304C;&#x540C;&#x5B9A;&#x3055;&#x308C;&#x3066;&#x3044;&#x308B;&#x3002;

&#x30A8;&#x30AF;&#x30B9;&#x30D1;&#x30F3;&#x30B7;&#x30F3;&#x306E;&#x6210;&#x9577;&#x4E16;&#x754C;

Expansins&#xB294; &#xD2B9;&#xC131; &#xD558; &#xACE0; &#xB3C5;&#xD2B9;&#xD55C; &#xBC29;&#xC2DD;&#xC73C;&#xB85C; Ph&#xC5D0; &#xC885;&#xC18D;&#xC801;&#xC778; &#xBCBD; &#xD655;&#xC7A5; &#xBC0F; &#xC2A4;&#xD2B8;&#xB808;&#xC2A4; &#xC774;&#xC644;&#xC744; &#xC720;&#xB3C4; &#xD558;&#xB294; &#xC138;&#xD3EC; &#xBCBD; &#xB2E8;&#xBC31;&#xC9C8;. &#xC54C;&#xD30C; &#xBC0F; &#xBCA0;&#xD0C0; expansins, &#xB77C;&#xB294; expansins&#xC758; &#xB450; &#xAC00;&#xC871; &#xC54C;&#xB824;&#xC838; &#xC788;&#xB2E4; &#xADF8;&#xB9AC;&#xACE0; &#xADF8;&#xB4E4;&#xC740; &#xD070; multigene &#xAC00;&#xC871; &#xAD6C;&#xC131;&#xC6D0;&#xC774; &#xBCF4;&#xAE30; &#xB2E4;&#xC591; &#xD55C; &#xC624;&#xB974;&#xAC04;-, &#xC870;&#xC9C1; &#xBC0F; &#xC138;&#xD3EC; &#xAD00;&#xB828; &#xC2DD; &#xD328;&#xD134;&#xC744; &#xAD6C;&#xC131; &#xD558;. Expansins &#xBA3C; &#xC2DC;&#xD000;&#xC2A4; &#xC720;&#xC0AC;&#xC131; &#xACF0; &#xB2E4;&#xB978; &#xC720;&#xC804;&#xC790; &#xC2DC;&#xD000;&#xC2A4; &#xB370;&#xC774;&#xD130;&#xBCA0;&#xC774;&#xC2A4;&#xC5D0; &#xD45C;&#xC2DC;&#xB3C4; &#xB429;&#xB2C8;&#xB2E4; &#xD558;&#xC9C0;&#xB9CC; &#xADF8;&#xB4E4;&#xC758; &#xC0DD;&#xBB3C; &#xD559;&#xC801; &#xBC0F; &#xC0DD; &#xD654; &#xD655; &#xC801;&#xC778; &#xAE30;&#xB2A5; &#xC544;&#xC9C1; &#xB418;&#xC9C0; &#xC54A;&#xC740; &#xBC1C;&#xACAC;. Expansin &#xAE00; &#xB8E8; &#xCE78;-&#xAE00; &#xB8E8; &#xCE78; &#xBC14;&#xC778;&#xB529;&#xC744; &#xC57D;&#xD558;&#xAC8C; &#xB098;&#xD0C0;&#xB098;&#xC9C0;&#xB9CC; &#xC791;&#xC5C5;&#xC758; &#xC790;&#xC138;&#xD55C; &#xBA54;&#xCEE4;&#xB2C8;&#xC998;&#xC740; &#xC798; &#xC124;&#xB9BD;. Expansins&#xC758; &#xC0DD;&#xBB3C;&#xD559;&#xC801; &#xC5ED;&#xD560; &#xB2E4;&#xC591; &#xD55C;, &#xD558;&#xC9C0;&#xB9CC; &#xC140; &#xD655;&#xB300;, &#xBD80;&#xB4DC;&#xB7FD;&#xAC8C; &#xD558;&#xB294; &#xACFC;&#xC77C;, &#xAF43;&#xAC00;&#xB8E8; &#xAD00; &#xBC0F; &#xB8E8;&#xD2B8; &#xD5E4;&#xC5B4; &#xC131;&#xC7A5; &#xBC0F; &#xC808;&#xB2E8; &#xD558;&#xB294; &#xB3D9;&#xC548; &#xC608;&#xB97C; &#xB4E4;&#xC5B4; &#xC138;&#xD3EC; &#xBCBD;&#xC744; &#xC644;&#xD654; &#xD558;&#xB294; expansins&#xC758; &#xD589;&#xB3D9;&#xC5D0; &#xAD00;&#xB828;&#xC774; &#xC788;&#xC744; &#xC218; &#xC788;&#xC2B5;&#xB2C8;&#xB2E4;. Expansin &#xAC19;&#xC740; &#xB2E8;&#xBC31;&#xC9C8;&#xB3C4; &#xBC15;&#xD14C;&#xB9AC;&#xC544;&#xC640; &#xACF0; &#xD321;&#xC774;, &#xC5B4;&#xB514; &#xADF8;&#xB4E4;&#xC740; &#xACF5;&#xC7A5; &#xBCF8;&#xBB38;&#xC758; &#xBBF8;&#xC0DD;&#xBB3C; &#xCE68;&#xC785; &#xC218; &#xC788;&#xC2B5;&#xB2C8;&#xB2E4; &#xC6D0;&#xC870;&#xC5D0;&#xC11C; &#xD655;&#xC778; &#xB418;&#xC5C8;&#xC2B5;&#xB2C8;&#xB2E4;.

Expansins&#xC758; &#xC131;&#xC7A5; &#xD558; &#xACE0; &#xC788;&#xB294; &#xC138;&#xACC4;.

Expansinas son prote&#xED;nas de la pared celular que inducen dependiente del pH de extensi&#xF3;n de la pared y relajaci&#xF3;n de la tensi&#xF3;n de una manera caracter&#xED;stica y &#xFA;nica. Dos familias de expansinas son conocidos, llamado alfa y beta expansinas, y comprenden las familias numerosas cuyos miembros multigene muestran diversos &#xF3;rganos, tejidos y c&#xE9;lulas espec&#xED;ficas de los patrones de expresi&#xF3;n. Otros genes que llevan a la similitud de secuencias distantes expansinas tambi&#xE9;n est&#xE1;n representados en las bases de datos de secuencias, pero sus funciones biol&#xF3;gicas y bioqu&#xED;micas a&#xFA;n no han sido descubiertas. Expansina parece debilitar glucano-glucano vinculante, pero su mecanismo de acci&#xF3;n detallado que no est&#xE1; bien establecida. Las funciones biol&#xF3;gicas de expansinas son diversos, pero puede estar relacionado con la acci&#xF3;n de expansinas para aflojar las paredes celulares, por ejemplo durante la ampliaci&#xF3;n de la c&#xE9;lula, ablandamiento del fruto, tubo de polen y el crecimiento del cabello de ra&#xED;z, y abscisi&#xF3;n. Expansina prote&#xED;nas similares tambi&#xE9;n se han identificado en bacterias y hongos, en los que pueden ayudar a la invasi&#xF3;n microbiana del cuerpo de la planta.

El creciente mundo de expansinas

Coenzyme Q(0) (Q(0)), a strong electrophile, is toxic to insulin-producing cells. Q(0) was incubated with rat and human pancreatic islets and INS-1 insulinoma cells, and its attachment to cellular proteins was studied with Western analysis using antiserum raised against the benzoquinone ring structure of ubiquinone (anti-Q). Q(0) covalently bonded to two proteins, one of 50 kDa and another of 70 kDa. Both proteins were found to be mitochondrial in human and rat islet cells and in many rat organs. Mitochondria were incubated with Q(0), and affinity-purified anti-Q was used to immunoprecipitate the 50-kDa protein. Amino acid sequencing identified it as dihydrolipoamide succinyltransferase, the E2 component of the alpha-ketoglutarate dehydrogenase complex (KDC). Western analysis also showed that Q bonds to the E2 components of the purified KDC and (0)the pyruvate dehydrogenase complex (PDC). Dihydrolipoamide acetyltransferase, the E2 of the PDC, has a molecular mass of 70 kDa, and the 70-kDa protein was inferred to be this enzyme. Q(0) was found to bond only to proteins containing dihydrolipoate, and in preparations of mitochondria, thiol reducing agents facilitated the attachment of Q(0), but oxidizing agents prevented it, suggesting that Q(0) bonds to thiols of dihydrolipoamide. Incubation of human or pig PDC with Q(0) followed by matrix-assisted laser desorption ionization time-of-flight and liquid chromatography/electrospray ionization mass spectrometry analyses of chymotrypsin-digested peptides of PDC E2 confirmed that Q(0) bonds to the dihydrolipoamide in these proteins. In mitochondria, coenzymes Q(1) and Q(2) did not bond to the 50-kDa protein but competed with the bonding of Q(0) to this protein. The prevention by Q(1) of characteristics the bonding of Q(0) to KDC E2, as well as other of the Q(0) effect, are reminiscent of the action of Q(0) on the mitochondrial permeability transition pore described previously (Fontaine, E., Ichas, F., and Bernardi, P. (1998) J. Biol. Chem. 273, 25734-25740).

Immunochemical identification of coenzyme Q0-dihydrolipoamide adducts in the E2 components of the alpha-ketoglutarate and pyruvate dehydrogenase complexes partially explains the cellular toxicity of coenzyme Q0.

UDP sulfoquinovose &#x5408;&#x9176;&#xFF0C;SQD1&#xFF0C;&#x50AC;&#x5316;&#x4E9A;&#x786B;&#x9178;&#x76D0;&#x79FB;&#x4EA4; UDP sulfoquinovose&#xFF0C;&#x8FD9;&#x662F;&#x4E3A;&#x751F;&#x7269;&#x5408;&#x6210;&#x7684;&#x690D;&#x7269; sulfolipid sulfoquinovosyldiacylglyerol &#x5934;&#x96C6;&#x56E2;&#x6350;&#x52A9; UDP-&#x8461;&#x8404;&#x7CD6;&#x5F15;&#x8D77;&#x3002;&#x83E0;&#x83DC;&#x672C;&#x673A; SQD1 &#x9176;&#x4F5C;&#x4E3A; 250 kDa heteroprotein &#x590D;&#x6742;&#x591A;&#x4EB2;&#x548C;&#x529B;&#x4E3A;&#x886C;&#x5E95;&#x4E9A;&#x786B;&#x9178;&#x76D0;&#x8981;&#x9AD8;&#x4E8E;&#x91CD;&#x7EC4; SQD1 &#x86CB;&#x767D;&#x8D28;&#x672C;&#x8EAB;&#x7684;&#x5B58;&#x5728;&#x3002;SQD1 &#x86CB;&#x767D; co-purified &#x4E0E;&#x4E5D;&#x4E2A;&#x86CB;&#x767D;&#x8D28;&#x3002;&#x53EF;&#x80FD;&#x7ED1;&#x5B9A;&#x4F19;&#x4F34;&#x5305;&#x62EC;&#x5149;&#x5408;&#x4F5C;&#x7528;&#x3001; &#x70ED;&#x4F11;&#x514B;&#x86CB;&#x767D; 70 &#x548C;&#x94C1;&#x6C27;&#x8FD8;&#x86CB;&#x767D;&#x4F9D;&#x8D56;&#x8C37;&#x6C28;&#x9178;&#x5408;&#x9176; (FdGOGAT)&#x3002;&#x867D;&#x7136;&#x5DF2;&#x77E5;&#x7684;&#x524D;&#x4E24;&#x4E2A;&#x86CB;&#x767D;&#x8D28;&#x4E0E;&#x8BB8;&#x591A;&#x5176;&#x4ED6;&#x86CB;&#x767D;&#x8D28;&#x8FDB;&#x884C;&#x4EA4;&#x4E92;&#xFF0C;FdGOGAT &#x7684;&#x9274;&#x5B9A;&#x662F;&#x6700;&#x8FF7;&#x4EBA;&#x7684;&#x56E0;&#x4E3A;&#x6B64; 160kDa &#x86CB;&#x767D;&#x5305;&#x542B;&#x5DF2;&#x77E5;&#x7ED1;&#x5B9A;&#x4E9A;&#x786B;&#x9178;&#x4F53;&#x5916; FMN &#x8F85;&#x9176;&#x3002;&#x4F7F;&#x7528;&#x4E0D;&#x540C;&#x7684;&#x6784;&#x9020;&#xFF0C;&#x91CD;&#x7EC4;&#x7684;&#x8868;&#x8FBE;&#x5F62;&#x5F0F;&#x591A;&#x57DF;&#x86CB;&#x767D; FdGOGAT&#xFF0C;&#x5B83;&#x8868;&#x660E; FMN &#x7ED1;&#x5B9A;&#x57DF;&#x7684; FdGOGAT &#x662F;&#x5230; SQD1 &#x86CB;&#x767D;&#x7684;&#x7279;&#x5F02;&#x6027;&#x7ED3;&#x5408;&#x7684;&#x5FC5;&#x8981;&#x6761;&#x4EF6;&#x3002;&#x4E00;&#x4E2A;&#x6A21;&#x578B;&#x8868;&#x660E; FdGOGAT &#x53EF;&#x4EE5;&#x901A;&#x9053;&#x5230; SQD1 &#x7684;&#x4E9A;&#x786B;&#x9178;&#x76D0;&#x3002;

&#x94C1;&#x6C27;&#x8FD8;&#x86CB;&#x767D;&#x4F9D;&#x8D56;&#x8C37;&#x6C28;&#x9178;&#x5408;&#x9176;&#x6708;&#x5149;&#x4E2D;&#x690D;&#x7269; sulfolipid &#x751F;&#x7269;&#x5408;&#x6210;&#x7684;&#x5F62;&#x6210;&#x4E0E; SQD1 &#x7684;&#x590D;&#x6742;&#x3002;

UDP-sulfoquinovose synthase, SQD1, catalyzes the transfer of sulfite to UDP-glucose giving rise to UDP-sulfoquinovose, which is the head group donor for the biosynthesis of the plant sulfolipid sulfoquinovosyldiacylglyerol. The native SQD1 enzyme of spinach exists as a 250 kDa heteroprotein complex with much higher affinity for the substrate sulfite than the recombinant SQD1 protein itself. The SQD1 protein co-purified with nine proteins. Likely binding partners included rubisco activase, HSP70, and ferredoxin-dependent glutamate synthase (FdGOGAT). While the first two proteins are known to interact with many other proteins, the identification of FdGOGAT was most intriguing because this 160kDa protein contains an FMN cofactor known to bind sulfite in vitro. Using different constructs expressing recombinant forms of the multidomain protein FdGOGAT, it was demonstrated that the FMN-binding domain of FdGOGAT is essential for specific binding of the protein to SQD1. A model suggests that FdGOGAT could channel sulfite to SQD1.

Ferredoxin-dependent glutamate synthase moonlights in plant sulfolipid biosynthesis by forming a complex with SQD1.

UDP-sulfoquinovose synthase, SQD1, catalyse le transfert du sulfite d&#x26;#39;UDP-glucose donnant lieu &#xE0; l&#x26;#39;UDP-sulfoquinovose, qui est le donneur de groupe de t&#xEA;te pour la biosynth&#xE8;se de l&#x26;#39;sulfoquinovosyldiacylglyerol sulfolipide plante. Le natif de SQD1 enzyme d&#x26;#39;&#xE9;pinards existe comme un complexe 250 kDa h&#xE9;t&#xE9;roprot&#xE9;ine avec une affinit&#xE9; beaucoup plus &#xE9;lev&#xE9;e pour le substrat que le sulfite recombinant SQD1 prot&#xE9;ine elle-m&#xEA;me. Le SQD1 prot&#xE9;ine co-purifi&#xE9; avec neuf prot&#xE9;ines. Probables partenaires de liaison inclus rubisco activase, HSP70, et glutamate synthase ferr&#xE9;doxine-d&#xE9;pendante (FdGOGAT). Alors que les deux premi&#xE8;res prot&#xE9;ines sont connues pour interagir avec de nombreuses autres prot&#xE9;ines, l&#x26;#39;identification des FdGOGAT &#xE9;tait la plus intrigante, car cette prot&#xE9;ine 160 kDa contient un cofacteur FMN connue pour se lier in vitro sulfite. En utilisant diff&#xE9;rentes constructions exprimant des formes recombinantes de la prot&#xE9;ine multidomaine FdGOGAT, il a &#xE9;t&#xE9; d&#xE9;montr&#xE9; que le domaine FMN contraignant de FdGOGAT est essentiel pour la liaison sp&#xE9;cifique de la prot&#xE9;ine &#xE0; SQD1. Un mod&#xE8;le sugg&#xE8;re que FdGOGAT pourrait canal sulfite &#xE0; SQD1.

Ferr&#xE9;doxine-d&#xE9;pendants Moonlights glutamate synthase dans la biosynth&#xE8;se sulfolipide usine en formant un complexe avec SQD1

Ferredoxin-abh&#xE4;ngige Glutamat-Synthase Mondlicht in Pflanzenzellen Sulfolipid-Biosynthese durch Bildung eines Komplexes mit SQD1.

Sintasi UDP-sulfoquinovose, SQD1, catalizza il trasferimento di solfito che d&agrave; origine a UDP-sulfoquinovose, che &egrave; il capo gruppo donatore per la biosintesi della pianta sulfolipid sulfoquinovosyldiacylglyerol UDP-glucosio. L&#39;enzima SQD1 nativo di spinaci esiste come un 250 kDa heteroprotein complesso con molta maggiore affinit&agrave; per il solfito di substrato di recombinant della proteina di SQD1 stesso. La proteina SQD1 co-purified con nove proteine. Partner associazione probabilmente incluso activase di rubisco, HSP70 e ferredossina-dipendente glutammato sintasi (FdGOGAT). Mentre le prime due proteine sono noti per interagire con molte altre proteine, l&#39;identificazione del FdGOGAT era pi&ugrave; intrigante perch&eacute; questa proteina 160kDa contiene un cofattore FMN noto per associare il solfito in vitro. Utilizzando diversi costrutti che esprimono forme recombinant della proteina multidominio FdGOGAT, &egrave; stato dimostrato che il dominio FMN-associazione di FdGOGAT &egrave; essenziale per l&#39;associazione specifica della proteina di SQD1. Un modello suggerisce che FdGOGAT potrebbe canale solfito a SQD1.

Ferredossina-dipendente glutammato sintasi lune nella biosintesi sulfolipid pianta formando un complesso con SQD1.

UDP-sulfoquinovose&#x30B7;&#x30F3;&#x30BF;&#x30FC;&#x30BC;&#x3001;SQD1&#x306F;&#x3001;&#x690D;&#x7269;&#x30B9;&#x30EB;&#x30DB;&#x30EA;&#x30D4;&#x30C9;&#x306E;sulfoquinovosyldiacylglyerol&#x306E;&#x751F;&#x5408;&#x6210;&#x306E;&#x305F;&#x3081;&#x306E;&#x982D;&#x90E8;&#x57FA;&#x4F9B;&#x4E0E;&#x4F53;&#x3067;&#x3042;&#x308B;UDP-sulfoquinovose&#x3001;&#x3092;&#x751F;&#x307F;&#x51FA;&#x3057;&#x3066;UDP-&#x30B0;&#x30EB;&#x30B3;&#x30FC;&#x30B9;&#x306B;&#x4E9C;&#x786B;&#x9178;&#x5869;&#x306E;&#x8EE2;&#x79FB;&#x3092;&#x89E6;&#x5A92;&#x3057;&#x307E;&#x3059;&#x3002;&#x307B;&#x3046;&#x308C;&#x3093;&#x8349;&#x306E;&#x30CD;&#x30A4;&#x30C6;&#x30A3;&#x30D6;SQD1&#x9175;&#x7D20;&#x306F;&#x3001;&#x7D44;&#x63DB;&#x3048;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;SQD1&#x81EA;&#x4F53;&#x3088;&#x308A;&#x4E9C;&#x786B;&#x9178;&#x57FA;&#x677F;&#x306E;&#x306F;&#x308B;&#x304B;&#x306B;&#x9AD8;&#x3044;&#x89AA;&#x548C;&#x6027;&#x3092;&#x6301;&#x3064;250 kDa&#x306E;heteroprotein&#x8907;&#x5408;&#x4F53;&#x3068;&#x3057;&#x3066;&#x5B58;&#x5728;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002; SQD1&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;9&#x86CB;&#x767D;&#x8CEA;&#x3068;&#x5171;&#x7CBE;&#x88FD;&#x3055;&#x308C;&#x305F;&#x3002;&#x304A;&#x305D;&#x3089;&#x304F;&#x7D50;&#x5408;&#x30D1;&#x30FC;&#x30C8;&#x30CA;&#x30FC;&#x306F;&#x3001;HSP70&#x30EB;&#x30D3;&#x30B9;&#x30B3;&#x30A2;&#x30AF;&#x30C1;&#x30D0;&#x30FC;&#x30BC;&#x3001;&#x304A;&#x3088;&#x3073;&#x30D5;&#x30A7;&#x30EC;&#x30C9;&#x30AD;&#x200B;&#x200B;&#x30B7;&#x30F3;&#x4F9D;&#x5B58;&#x6027;&#x30B0;&#x30EB;&#x30BF;&#x30DF;&#x30F3;&#x9178;&#x5408;&#x6210;&#x9175;&#x7D20;&#xFF08;FdGOGAT&#xFF09;&#x304C;&#x542B;&#x307E;&#x308C;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x6700;&#x521D;&#x306E;&#x4E8C;&#x3064;&#x306E;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x306F;&#x4ED6;&#x306E;&#x591A;&#x304F;&#x306E;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x3068;&#x76F8;&#x4E92;&#x4F5C;&#x7528;&#x3059;&#x308B;&#x3053;&#x3068;&#x304C;&#x77E5;&#x3089;&#x308C;&#x3066;&#x3044;&#x308B;&#x304C;&#x3001;&#x3053;&#x306E;160kDa&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x304C;in vitro&#x3067;&#x4E9C;&#x786B;&#x9178;&#x5869;&#x7D50;&#x5408;&#x3059;&#x308B;&#x3053;&#x3068;&#x304C;&#x77E5;&#x3089;&#x308C;&#x3066;FMN&#x88DC;&#x56E0;&#x5B50;&#x304C;&#x542B;&#x307E;&#x308C;&#x3066;&#x3044;&#x308B;&#x305F;&#x3081;&#x3001;FdGOGAT&#x306E;&#x540C;&#x5B9A;&#x304C;&#x6700;&#x3082;&#x8208;&#x5473;&#x3092;&#x305D;&#x305D;&#x3089;&#x306A;&#x304B;&#x3063;&#x305F;&#x3002;&#x30DE;&#x30EB;&#x30C1;&#x30C9;&#x30E1;&#x30A4;&#x30F3;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;FdGOGAT&#x306E;&#x7D44;&#x63DB;&#x3048;&#x5F62;&#x614B;&#x3092;&#x8868;&#x73FE;&#x3059;&#x308B;&#x5225;&#x306E;&#x69CB;&#x6587;&#x3092;&#x4F7F;&#x7528;&#x3057;&#x3066;&#x3001;&#x305D;&#x308C;&#x306F;FdGOGAT&#x306E;FMN&#x7D50;&#x5408;&#x30C9;&#x30E1;&#x30A4;&#x30F3;&#x306F;SQD1&#x3078;&#x306E;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x306E;&#x7279;&#x7570;&#x7684;&#x7D50;&#x5408;&#x306B;&#x5FC5;&#x9808;&#x3067;&#x3042;&#x308B;&#x3053;&#x3068;&#x304C;&#x793A;&#x3055;&#x308C;&#x305F;&#x3002;&#x30E2;&#x30C7;&#x30EB;&#x306F;FdGOGAT&#x304C;&#x30C1;&#x30E3;&#x30CD;&#x30EB;SQD1&#x306B;&#x4E9C;&#x786B;&#x9178;&#x3053;&#x3068;&#x3092;&#x793A;&#x5506;&#x3057;&#x3066;&#x3044;&#x308B;&#x3002;

SQD1&#x3068;&#x8907;&#x5408;&#x4F53;&#x3092;&#x5F62;&#x6210;&#x3059;&#x308B;&#x3053;&#x3068;&#x306B;&#x3088;&#x308A;&#x690D;&#x7269;&#x30B9;&#x30EB;&#x30DB;&#x30EA;&#x30D4;&#x30C9;&#x751F;&#x5408;&#x6210;&#x306B;&#x304A;&#x3051;&#x308B;&#x30D5;&#x30A7;&#x30EC;&#x30C9;&#x30AD;&#x200B;&#x200B;&#x30B7;&#x30F3;&#x4F9D;&#x5B58;&#x6027;&#x30B0;&#x30EB;&#x30BF;&#x30DF;&#x30F3;&#x9178;&#x5408;&#x6210;&#x9175;&#x7D20;&#x6708;&#x5149;

UDP-sulfoquinovose synthase, SQD1, &#xD669;&#xC0B0;&#xC758; UDP-sulfoquinovose, &#xACF5;&#xC7A5; sulfolipid sulfoquinovosyldiacylglyerol&#xC758; &#xC0DD; &#xD569;&#xC131;&#xC5D0; &#xB300; &#xD55C; &#xBA38;&#xB9AC; &#xADF8;&#xB8F9; &#xAE30;&#xC99D;&#xC790;&#xB294; UDP-&#xD3EC;&#xB3C4; &#xB2F9; &#xC8FC;&#xB294; &#xC0C1;&#xC2B9; &#xC804;&#xC1A1;&#xC744; catalyzes. &#xC2DC;&#xAE08;&#xCE58;&#xC758; &#xB124;&#xC774;&#xD2F0;&#xBE0C; SQD1 &#xD6A8;&#xC18C;&#xB97C; 250 kDa heteroprotein &#xBCF5;&#xC7A1; &#xD55C; &#xC7AC;&#xC870;&#xD569; SQD1 &#xB2E8;&#xBC31;&#xC9C8; &#xC790;&#xCCB4; &#xBCF4;&#xB2E4; &#xAE30;&#xD310; &#xD669;&#xC0B0;&#xC5D0; &#xB300; &#xD55C; &#xD6E8;&#xC52C; &#xB192;&#xC740; &#xC120;&#xD638;&#xB3C4; &#xC874;&#xC7AC;&#xD569;&#xB2C8;&#xB2E4;. 9 &#xAC1C;&#xC758; &#xB2E8;&#xBC31;&#xC9C8; co-purified SQD1 &#xB2E8;&#xBC31;&#xC9C8;&#xC785;&#xB2C8;&#xB2E4;. &#xAC00;&#xB2A5;&#xC131;&#xC774; &#xBC14;&#xC778;&#xB529; &#xD30C;&#xD2B8;&#xB108; rubisco activase, HSP70, ferredoxin &#xC885;&#xC18D; &#xC0B0; &#xC5FC; synthase (FdGOGAT)&#xB97C; &#xD3EC;&#xD568; &#xD569;&#xB2C8;&#xB2E4;. &#xCC98;&#xC74C; &#xB450; &#xB2E8;&#xBC31;&#xC9C8; &#xB9CE;&#xC740; &#xB2E4;&#xB978; &#xB2E8;&#xBC31;&#xC9C8; &#xC0C1;&#xD638; &#xC791;&#xC6A9; &#xC54C;&#xB824;&#xC838; &#xC788;&#xC2B5;&#xB2C8;&#xB2E4;, 160kDa &#xB2E8;&#xBC31;&#xC9C8;&#xC774; &#xC544; &#xCCB4; &#xC678;&#xC5D0; &#xBC14;&#xC778;&#xB529;&#xD560; &#xC54C;&#xB824;&#xC9C4; FMN &#xACF5;&#xB3D9; &#xC778;&#xC790;&#xB97C; &#xD3EC;&#xD568; &#xD558;&#xAE30; &#xB54C;&#xBB38;&#xC5D0; Fdgogat&#xC758; &#xC2DD;&#xBCC4; &#xAC00;&#xC7A5; &#xD765;&#xBBF8; &#xD588;&#xB2E4;. &#xB2E4;&#xC911; &#xB3C4;&#xBA54;&#xC778; &#xB2E8;&#xBC31;&#xC9C8; Fdgogat&#xC758; &#xC7AC;&#xC870;&#xD569; &#xD615;&#xD0DC;&#xB97C; &#xD45C;&#xD604; &#xD558;&#xB294; &#xB2E4;&#xB978; &#xAD6C;&#xBB38;&#xC744; &#xC0AC;&#xC6A9; &#xD558; &#xC5EC;, FdGOGAT FMN &#xBC14;&#xC778;&#xB529; &#xB3C4;&#xBA54;&#xC778; SQD1 &#xB2E8;&#xBC31;&#xC9C8;&#xC758; &#xD2B9;&#xC815; &#xBC14;&#xC778;&#xB529;&#xC5D0; &#xD544;&#xC218;&#xC801;&#xC785;&#xB2C8;&#xB2E4; &#xC2DC;&#xC5F0; &#xB418;&#xC5C8;&#xB2E4;. &#xBAA8;&#xB378; FdGOGAT SQD1 &#xD558; &#xC544; &#xCC44;&#xB110; &#xC218; &#xB098;&#xC654;&#xB2E4;.

&#xBC24;&#xC5D0; &#xBD80;&#xC5C5; ferredoxin &#xC885;&#xC18D; &#xC0B0; &#xC5FC; synthase&#xB85C; &#xBCF5;&#xC7A1; &#xD55C; SQD1 &#xD615;&#xC131; &#xD558; &#xC5EC; &#xACF5;&#xC7A5; sulfolipid &#xC0DD; &#xD569;&#xC131;&#xC5D0;.

UDP-sulfoquinovose sintasa, SQD1, cataliza la transferencia de sulfito a UDP-glucosa dando lugar a la UDP-sulfoquinovose, que es el donante grupo de cabeza de la bios&#xED;ntesis de la sulfoquinovosyldiacylglyerol sulfol&#xED;pidos planta. El nativo SQD1 enzima de espinaca existe como un complejo kDa 250 heteroprotein con una afinidad mucho mayor para el sustrato que el sulfito recombinante SQD1 propia prote&#xED;na. La prote&#xED;na SQD1 co-purifica con nueve prote&#xED;nas. Posibles socios vinculantes incluyen Rubisco activase, HSP70, y dependiente de ferredoxina glutamato sintasa (FdGOGAT). Mientras que los primeros dos prote&#xED;nas se sabe que interact&#xFA;an con muchas otras prote&#xED;nas, la identificaci&#xF3;n de FdGOGAT era m&#xE1;s intrigante, porque esta prote&#xED;na 160kDa contiene un cofactor FMN sabe que se unen al sulfito in vitro. Utilizando diferentes construcciones que expresan formas recombinantes de la prote&#xED;na FdGOGAT multidominio, se demostr&#xF3; que el dominio de uni&#xF3;n de FMN FdGOGAT es esencial para la uni&#xF3;n espec&#xED;fica de la prote&#xED;na a SQD1. Un modelo sugiere que podr&#xED;a FdGOGAT canal de sulfito de SQD1.

Ferredoxina dependientes de glutamato sintasa de Luz de luna en la bios&#xED;ntesis de la planta sulfol&#xED;pidos formando un complejo con SQD1

Post-translational modification of proteins is a dynamic way of generating new protein-protein interaction interfaces that are critical for signaling networks in diverse cellular functions. Purified recombinant proteins frequently lack these signature modifications. Using the tumor suppressor p53 as the model protein, we present here a tethered catalysis approach for the production of acetylated p53 in vivo. P53 is a major tumor suppressor protein that protects the cell from various oncogenic stresses. Upon DNA damage, p53 is stabilized and activated by a plethora of post-translational modifications, including acetylation. Here, we show that constitutively acetylated p53 can be expressed and purified from both yeast and Escherichia coli. This method is highly suitable for studying protein-protein interactions in the conventional yeast two-hybrid screen that requires a constitutively acetylated state of p53. Furthermore, effective production and purification of acetylated p53 from E. coli supports future biochemical and structural characterization. The method described in this work can be applied to other proteins and modifications, and thus has widespread use in the fields of signal transduction and proteomic research.

Production of constitutively acetylated recombinant p53 from yeast and Escherichia coli by tethered catalysis.

In budding yeast, the meiosis-specific protein kinase Ime2 is required for normal meiotic progression. Current evidence suggests that Ime2 is functionally related to Cdc28, the major cyclin-dependent kinase in yeast that is essential for both cell cycle and meiosis. We have previously reported that a natural target of Ime2 activity is replication protein A (RPA), the cellular single-stranded DNA-binding protein that performs critical functions during DNA replication, repair and recombination. Ime2-dependent RPA phosphorylation first occurs early in meiosis and targets the middle subunit of the RPA heterotrimeric complex (Rfa2). We now demonstrate that Rfa2 serine 27 (S27) is required for Ime2-dependent Rfa2 phosphorylation in vivo. S27 is also required for Rfa2 phosphorylation in vitro catalyzed by immunoprecipitated Ime2. In addition, Ime2 mediates in vitro phosphorylation of a short peptide containing Rfa2 amino acids 23 through 29, thereby providing evidence that S27 itself is the phosphoacceptor. Phosphorylation site mapping supports this conclusion, as mass spectrometry analysis has revealed that at least three residues within Rfa2 amino acids 2 through 35 become phosphorylated specifically during meiosis. Although S27 is embedded in a motif that is recognized by several protein kinases, this sequence is not a typical target of cyclin-dependent kinases. Therefore, the mechanism underlying Ime2 substrate recognition could differ from that of Cdc28.

Mechanistic insight into the Cdc28-related protein kinase Ime2 through analysis of replication protein A phosphorylation.

&#x79CD;&#x5B50;&#x53D1;&#x80B2;&#x8FC7;&#x7A0B;&#x4E2D;&#x7684;&#x5B58;&#x50A8;&#x590D;&#x5408;&#x79EF;&#x7D2F;&#x51C6;&#x5907;&#x690D;&#x7269;&#x751F;&#x5B58;&#x7684;&#x4E0B;&#x4E00;&#x4EE3;&#x3002;&#x56E0;&#x6B64;&#xFF0C;&#x8FC7;&#x7A0B;&#x53C2;&#x4E0E;&#x7684;&#x89C4;&#x7BA1;&#x548C;&#x5B58;&#x50A8;&#x590D;&#x5408;&#x5806;&#x79EF;&#x5728;&#x79CD;&#x5B50;&#x840C;&#x53D1;&#x548C;&#x5E7C;&#x82D7;&#x7684;&#x5EFA;&#x7ACB;&#x53D1;&#x5C55;&#x718A;&#x606F;&#x606F;&#x76F8;&#x5173;&#x7684;&#x5408;&#x6210;&#x3002;Wrinkled1 (wri1) &#x7A81;&#x53D8;&#x4F53;&#x7684;&#x62DF;&#x5357;&#x82A5; (&#x62DF;&#x5357;&#x82A5;) &#x6709;&#x635F;&#x4E8E;&#x79CD;&#x5B50;&#x6CB9;&#x79EF;&#x7D2F;&#x3002;WRI1 &#x57FA;&#x56E0;&#x7F16;&#x7801; APETALA2/&#x4E59;&#x70EF;&#x53CD;&#x5E94;&#x7684;&#x5143;&#x7D20;&#x7ED3;&#x5408;&#x86CB;&#x767D;&#x8F6C;&#x5F55;&#x56E0;&#x5B50;&#x53C2;&#x4E0E;&#x63A7;&#x5236;&#x7684;&#x4EE3;&#x8C22;&#xFF0C;&#x5C24;&#x5176;&#x662F;&#x7CD6;&#x9175;&#x89E3;&#xFF0C;&#x53D1;&#x5C55;&#x4E2D;&#x56FD;&#x5BB6;&#x7684;&#x79CD;&#x5B50;&#x7684;&#x3002;&#x5728;&#x8FD9;&#x91CC;&#x6211;&#x4EEC;&#x901A;&#x8FC7;&#x6BD4;&#x8F83;&#x8868;&#x793A;&#x4E2D;&#x7684; wri1-1 &#x7A81;&#x53D8;&#x4F53;&#x7684;&#x80CC;&#x666F;&#x548C;&#x91CE;&#x751F;&#x578B; WRI1 &#x91CE;&#x751F;&#x578B;&#x57FA;&#x56E0;&#x7684;&#x8F6C;&#x57FA;&#x56E0;&#x682A;&#x7684; wri1-1 &#x7A81;&#x53D8;&#x4F53;&#xFF0C;&#x8C03;&#x67E5;&#x4E2D;&#x79CD;&#x5B50;&#x840C;&#x53D1;&#x548C;&#x5E7C;&#x82D7;&#x5EFA;&#x7ACB;&#x6B64;&#x89C4;&#x7BA1;&#x56E0;&#x5B50;&#x7684;&#x4F5C;&#x7528;&#x3002;&#x690D;&#x7269;&#x4E2D; WRI1 &#x57FA;&#x56E0;&#x7684;&#x8868;&#x8FBE;&#x6539;&#x53D8;&#x663E;&#x793A;&#x4E0D;&#x540C;&#x840C;&#x53D1;&#x751F;&#x957F;&#x56E0;&#x5B50;&#x8131;&#x843D;&#x9178; (ABA)&#x3001; &#x7CD6;&#x7C7B;&#x548C;&#x8102;&#x80AA;&#x9178;&#x4ECB;&#x8D28;&#x4E2D;&#x63D0;&#x4F9B;&#x7684;&#x56DE;&#x5E94;&#x3002;&#x7A81;&#x53D8;&#x4F53;&#x840C;&#x53D1;&#x4E86;&#x5BF9;&#x963F;&#x575D;&#x3001; &#x7CD6;&#x7C7B;&#x548C; osmolites&#xFF0C;&#x51CF;&#x8F7B;&#x4E86;&#x8F6C;&#x57FA;&#x56E0;&#x682A;&#x589E;&#x52A0; WRI1 &#x8868;&#x8FBE;&#x7684;&#x5F71;&#x54CD;&#x66F4;&#x52A0;&#x654F;&#x611F;&#x3002;Wri1-1 &#x7A81;&#x53D8;&#x4F53;&#x4E2D;&#x589E;&#x52A0;&#x4E86;&#x57FA;&#x56E0;&#x8868;&#x8FBE;&#x7684;&#x963F;&#x575D;&#x54CD;&#x5E94; AtEM6 &#x548C; ABA &#x4E0D;&#x533A;&#x5206; 3 (ABI3)&#x3002;Abi3 3 &#x548C; wri1-1 &#x4E4B;&#x95F4;&#x7684;&#x53CC;&#x7A81;&#x53D8;&#x5206;&#x6790;&#x5EFA;&#x8BAE; WRI1 &#x548C; ABI3&#xFF0C;&#x8C03;&#x89E3;&#x79CD;&#x5B50;&#x3001; &#x963F;&#x575D;&#x53CD;&#x5E94;&#x7684;&#x8F6C;&#x5F55;&#x56E0;&#x5B50;&#x6CD5;&#x5728;&#x5E76;&#x884C;&#x7684;&#x9014;&#x5F84;&#x3002;&#x52A0;&#x6CD5; 2-&#x8131;&#x6C27;&#x8461;&#x8404;&#x7CD6;&#x6291;&#x5236;&#x79CD;&#x5B50;&#x840C;&#x53D1;&#xFF0C;&#x4F46;&#x5E76;&#x672A;&#x56E0;&#x6B64;&#x5C11;&#x8D85;&#x91CF;&#x8868;&#x8FBE; WRI1 &#x7684;&#x884C;&#x4E2D;&#x3002;&#x82D7; wri1-1 &#x7A81;&#x53D8;&#x4F53;&#x5728;&#x4E0B;&#x964D;&#xFF0C;&#x4F46;&#x80FD;&#x7F13;&#x89E3;&#x7684;&#x8517;&#x7CD6;&#x3002;&#x9664;&#x4E86;&#x840C;&#x53D1;&#x8FC7;&#x7A0B;&#x4E2D;&#x53EF;&#x80FD;&#x7684;&#x4FE1;&#x53F7;&#x4F5C;&#x7528;&#xFF0C;&#x4ECB;&#x8D28;&#x4E2D;&#x7684;&#x7CD6;&#x88AB;&#x8981;&#x6C42;&#x4F5C;&#x4E3A;&#x6784;&#x5EFA;&#x57FA;&#x5757;&#x548C;&#x80FD;&#x6E90;&#x4F9B;&#x5E94; wri1 1 &#x82D7;&#x5EFA;&#x7ACB;&#x8FC7;&#x7A0B;&#x3002;

WRI1 &#x662F;&#x5FC5;&#x9700;&#x7684;&#x79CD;&#x5B50;&#x840C;&#x53D1;&#x548C;&#x5E7C;&#x82D7;&#x7684;&#x5EFA;&#x7ACB;&#x3002;

Storage compound accumulation during seed development prepares the next generation of plants for survival. Therefore, processes involved in the regulation and synthesis of storage compound accumulation during seed development bear relevance to germination and seedling establishment. The wrinkled1 (wri1) mutant of Arabidopsis (Arabidopsis thaliana) is impaired in seed oil accumulation. The WRI1 gene encodes an APETALA2/ethylene-responsive element-binding protein transcription factor involved in the control of metabolism, particularly glycolysis, in the developing seeds. Here we investigate the role of this regulatory factor in seed germination and seedling establishment by comparing the wri1-1 mutant, transgenic lines expressing the WRI1 wild-type cDNA in the wri1-1 mutant background, and the wild type. Plants altered in the expression of the WRI1 gene showed different germination responses to the growth factor abscisic acid (ABA), sugars, and fatty acids provided in the medium. Germination of the mutant was more sensitive to ABA, sugars, and osmolites, an effect that was alleviated by increased WRI1 expression in transgenic lines. The expression of ABA-responsive genes AtEM6 and ABA-insensitive 3 (ABI3) was increased in the wri1-1 mutant. Double-mutant analysis between abi3-3 and wri1-1 suggested that WRI1 and ABI3, a transcription factor mediating ABA responses in seeds, act in parallel pathways. Addition of 2-deoxyglucose inhibited seed germination, but did so less in lines overexpressing WRI1. Seedling establishment was decreased in the wri1-1 mutant but could be alleviated by sucrose. Apart from a possible signaling role in germination, sugars in the medium were required as building blocks and energy supply during wri1-1 seedling establishment.

WRI1 is required for seed germination and seedling establishment.

L&#x26;#39;accumulation de compos&#xE9;s de stockage au cours du d&#xE9;veloppement des semences pr&#xE9;pare la prochaine g&#xE9;n&#xE9;ration de plantes pour survivre. Par cons&#xE9;quent, les processus impliqu&#xE9;s dans la r&#xE9;gulation et la synth&#xE8;se de l&#x26;#39;accumulation de compos&#xE9;s de stockage au cours du d&#xE9;veloppement des semences ours pertinence pour la germination et l&#x26;#39;&#xE9;tablissement des semis. Le WRINKLED1 (wri1) mutant d&#x26;#39;Arabidopsis (Arabidopsis thaliana) est alt&#xE9;r&#xE9;e dans l&#x26;#39;accumulation de l&#x26;#39;huile de graines. Le g&#xE8;ne code pour une WRI1 APETALA2/ethylene-responsive &#xE9;l&#xE9;ment de liaison de prot&#xE9;ine facteur de transcription impliqu&#xE9; dans la r&#xE9;gulation du m&#xE9;tabolisme, en particulier glycolyse, dans les graines de d&#xE9;veloppement. Nous &#xE9;tudions ici le r&#xF4;le de ce facteur de r&#xE9;gulation dans la germination des graines et l&#x26;#39;&#xE9;tablissement des semis en comparant les wri1-1 mutants, des lign&#xE9;es transg&#xE9;niques exprimant le WRI1 de type sauvage d&#x26;#39;ADNc dans le fond wri1-1 mutant, et le type sauvage. Plantes modifi&#xE9;es dans l&#x26;#39;expression du g&#xE8;ne WRI1 ont montr&#xE9; des r&#xE9;ponses diff&#xE9;rentes de germination &#xE0; l&#x26;#39;acide du facteur de croissance abscissique (ABA), de sucres et d&#x26;#39;acides gras fournis dans le milieu. La germination du mutant &#xE9;tait plus sensible &#xE0; l&#x26;#39;ABA, les sucres, et osmolites, un effet qui a &#xE9;t&#xE9; att&#xE9;nu&#xE9; par l&#x26;#39;augmentation WRI1 expression dans des lign&#xE9;es transg&#xE9;niques. L&#x26;#39;expression de l&#x26;#39;ABA-g&#xE8;nes sensibles AtEM6 et ABA-insensibles 3 (ABI3) a &#xE9;t&#xE9; augment&#xE9; dans le mutant wri1-1. Double-mutant analyse entre ABI3-3 et wri1-1 a sugg&#xE9;r&#xE9; que WRI1 et ABI3, un facteur de transcription la m&#xE9;diation des r&#xE9;ponses ABA dans les graines, acte dans des voies parall&#xE8;les. Ajout de 2-d&#xE9;soxyglucose inhibe la germination des graines, mais ont fait moins de lignes surexprimant WRI1. Mise en place des semis a &#xE9;t&#xE9; diminu&#xE9; dans le mutant wri1-1, mais pourrait &#xEA;tre att&#xE9;nu&#xE9;e par le saccharose. Mis &#xE0; part un r&#xF4;le de signalisation possible dans la germination, les sucres dans le milieu ont &#xE9;t&#xE9; tenus comme blocs de construction et l&#x26;#39;approvisionnement en &#xE9;nergie au cours de l&#x26;#39;&#xE9;tablissement wri1-1 plantule.

WRI1 est n&#xE9;cessaire &#xE0; la germination des semences et l&#x26;#39;&#xE9;tablissement des semis

Speicher zusammengesetzte Anh&auml;ufung w&auml;hrend der Samen Entwicklung bereitet die n&auml;chste Generation von Pflanzen f&uuml;r das &Uuml;berleben. Daher Prozesse beteiligt, in der Verordnung und die Synthese von Speicher zusammengesetzte Anh&auml;ufung w&auml;hrend der Samen Entwicklung B&auml;r Relevanz f&uuml;r Keimung und S&auml;mling Einrichtung. Die wrinkled1 (wri1)-Mutant von Arabidopsis (Arabidopsis Thaliana) Samen &Ouml;l Anh&auml;ufung beeintr&auml;chtigt ist. Das WRI1-Gen codiert f&uuml;r eine APETALA2/Ethylen-responsive Element-Binding Protein-&Uuml;bertragungfaktor haben, die Kontrolle des Stoffwechsels, insbesondere der Glykolyse, der entwickelnden Samen. Hier untersuchen wir die Rolle der dieser regulatorischen Faktor in Samen keimen und S&auml;mling Einrichtung durch den Vergleich der wri1-1 Mutant, transgener Linien WRI1 Wildtyp DNA in den wri1-1-mutant-Hintergrund und der Wildtyp zum Ausdruck zu bringen. Pflanzen, die in den Ausdruck des WRI1-Gens ver&auml;ndert zeigte verschiedene Keimung Antworten zu den Wachstumsfaktor Abscisic S&auml;ure (ABA), Zucker und Fetts&auml;uren, die im Medium bereitgestellt. Keimen von der Mutant war empfindlicher und ABA, Zucker und Osmolites, ein Effekt, der durch erh&ouml;hte WRI1-Expression in transgenen Linien gemildert wurde. Der Ausdruck der ABA-responsive Gene AtEM6 und ABA-unempfindliche 3 (ABI3) wurde in der wri1-1-Mutant erh&ouml;ht. Doppel-Mutant-Analyse von abi3-3 bis wri1-1 vorgeschlagen, WRI1 und ABI3, einen Transkriptionsfaktor Vermittlung ABA Antworten im Bereich Seeds handeln parallele Wege. Zusatz von 2-Deoxyglucose Samenkeimung gehemmt, aber tat also weniger in &uuml;berexprimierenden WRI1 Linien. S&auml;mling Niederlassung wurde in der wri1-1-Mutant zur&uuml;ckgegangen, sondern durch Saccharose gelindert werden konnte. Abgesehen von einer m&ouml;glichen Signalling Rolle bei Keimung waren Zucker in das Medium erforderlich, wie bei wri1-1 S&auml;mling Aufstellung Bausteine und Energie liefern.

WRI1 ist f&uuml;r Samen keimen und S&auml;mling Einrichtung erforderlich.

Accumulo di composto archiviazione durante lo sviluppo del seme si prepara la prossima generazione di piante per la sopravvivenza. Pertanto, i processi coinvolti nella sintesi di composti accumulo archiviazione durante la pertinenza di orso sviluppo seme germinazione e piantina istituzione e regolamento. Il wrinkled1 (wri1) mutante di Arabidopsis (Arabidopsis thaliana) &egrave; alterata in accumulo di olio di semi. Il WRI1 gene codifica per un fattore di trascrizione element-binding protein APETALA2/etilene-reattiva coinvolto nel controllo del metabolismo, particolarmente la glicolisi, nei semi in via di sviluppo. Qui indaghiamo il ruolo di questo fattore regolatore in stabilimento di germinazione e piantina di sementi confrontando il mutante di wri1-1, linee transgeniche che esprimono il cDNA wild-type WRI1 in background mutante wri1-1 e il tipo di selvatico. Piante modificate nell&#39;espressione del gene della WRI1 ha mostrato risposte germinazione differenti per il fattore di crescita acido abscissico (ABA), zuccheri e acidi grassi forniti nel mezzo. Germinazione del mutante &egrave; stato pi&ugrave; sensibile all&#39;ABA, zuccheri e osmolites, un effetto che &egrave; stato alleviato da aumentata espressione di WRI1 in linee transgeniche. L&#39;espressione dei geni ABA-responsive AtEM6 e ABA-insensibile 3 (ABI3) &egrave; stato aumentato in mutante wri1-1. Double-mutante analisi tra 3 abi3 e wri1-1 ha suggerito che WRI1 e ABI3, un fattore di trascrizione mediare risposte ABA in semi, agire percorsi in parallelo. Aggiunta di 2-deossiglucosio inibisce la germinazione dei semi, ma ha fatto cos&igrave; meno nelle linee con sovraesprimono relativa WRI1. Istituzione del semenzale era diminuito in mutante wri1-1, ma potrebbe essere alleviata da saccarosio. Oltre a un possibile ruolo di segnalazione in germinazione, zuccheri nel mezzo sono stati richiesti come blocchetti di costruzione e l&#39;energia di alimentazione durante l&#39;istituzione del semenzale di wri1-1.

WRI1 &egrave; richiesto per stabilimento di germinazione e piantina di sementi.

&#x7A2E;&#x5B50;&#x306E;&#x958B;&#x767A;&#x6642;&#x306B;&#x30B9;&#x30C8;&#x30EC;&#x30FC;&#x30B8;&#x5316;&#x5408;&#x7269;&#x306E;&#x84C4;&#x7A4D;&#x304C;&#x751F;&#x5B58;&#x306E;&#x305F;&#x3081;&#x306E;&#x6B21;&#x4E16;&#x4EE3;&#x306E;&#x690D;&#x7269;&#x3092;&#x6E96;&#x5099;&#x3057;&#x307E;&#x3059;&#x3002;&#x3057;&#x305F;&#x304C;&#x3063;&#x3066;&#x3001;&#x767A;&#x82BD;&#x306B;&#x7A2E;&#x5B50;&#x958B;&#x767A;&#x30AF;&#x30DE;&#x3068;&#x306E;&#x95A2;&#x9023;&#x6027;&#x3068;&#x82D7;&#x7ACB;&#x3061;&#x306E;&#x9593;&#x306B;&#x30B9;&#x30C8;&#x30EC;&#x30FC;&#x30B8;&#x5316;&#x5408;&#x7269;&#x306E;&#x84C4;&#x7A4D;&#x306E;&#x8ABF;&#x7BC0;&#x304A;&#x3088;&#x3073;&#x5408;&#x6210;&#x306B;&#x95A2;&#x4E0E;&#x3057;&#x3066;&#x51E6;&#x7406;&#x3057;&#x307E;&#x3059;&#x3002;&#x30B7;&#x30ED;&#x30A4;&#x30CC;&#x30CA;&#x30BA;&#x30CA;&#x306E;&#x5909;&#x7570;&#x4F53;wrinkled1&#xFF08;wri1&#xFF09;&#xFF08;&#x30B7;&#x30ED;&#x30A4;&#x30CC;&#x30CA;&#x30BA;&#x30CA;&#xFF09;&#x306E;&#x7A2E;&#x5B50;&#x6CB9;&#x306E;&#x84C4;&#x7A4D;&#x304C;&#x640D;&#x306A;&#x308F;&#x308C;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002; WRI1&#x907A;&#x4F1D;&#x5B50;&#x306F;&#x3001;&#x767A;&#x5C55;&#x9014;&#x4E0A;&#x306E;&#x7A2E;&#x5B50;&#x306B;&#x306F;&#x3001;&#x7279;&#x306B;&#x89E3;&#x7CD6;&#x4EE3;&#x8B1D;&#x306E;&#x5236;&#x5FA1;&#x306B;&#x95A2;&#x4E0E;APETALA2/ethylene-responsive&#x30A8;&#x30EC;&#x30E1;&#x30F3;&#x30C8;&#x7D50;&#x5408;&#x86CB;&#x767D;&#x8CEA;&#x306E;&#x8EE2;&#x5199;&#x56E0;&#x5B50;&#x3092;&#x30B3;&#x30FC;&#x30C9;&#x3057;&#x3066;&#x3044;&#x308B;&#x3002;&#x3053;&#x3053;&#x3067;&#x306F;wri1-1&#x5909;&#x7570;&#x4F53;&#x30D0;&#x30C3;&#x30AF;&#x30B0;&#x30E9;&#x30A6;&#x30F3;&#x30C9;&#x3067;WRI1&#x91CE;&#x751F;&#x578B;&#x306E;cDNA&#x3092;&#x767A;&#x73FE;&#x3057;&#x3066;&#x3044;&#x308B;wri1-1&#x5909;&#x7570;&#x4F53;&#x3001;&#x30C8;&#x30E9;&#x30F3;&#x30B9;&#x30B8;&#x30A7;&#x30CB;&#x30C3;&#x30AF;&#x7DDA;&#x3001;&#x304A;&#x3088;&#x3073;&#x91CE;&#x751F;&#x578B;&#x3092;&#x6BD4;&#x8F03;&#x3059;&#x200B;&#x200B;&#x308B;&#x3053;&#x3068;&#x306B;&#x3088;&#x3063;&#x3066;&#x3001;&#x7A2E;&#x5B50;&#x306E;&#x767A;&#x82BD;&#x3068;&#x82D7;&#x7ACB;&#x3061;&#x306B;&#x3053;&#x306E;&#x8ABF;&#x7BC0;&#x56E0;&#x5B50;&#x306E;&#x5F79;&#x5272;&#x3092;&#x8ABF;&#x3079;&#x308B;&#x3002; WRI1&#x907A;&#x4F1D;&#x5B50;&#x306E;&#x767A;&#x73FE;&#x306B;&#x5909;&#x66F4;&#x690D;&#x7269;&#x306F;&#x6210;&#x9577;&#x56E0;&#x5B50;&#x30A2;&#x30D6;&#x30B7;&#x30B8;&#x30F3;&#x9178;&#xFF08;ABA&#xFF09;&#x3001;&#x7CD6;&#x3001;&#x304A;&#x3088;&#x3073;&#x5A92;&#x4F53;&#x3067;&#x63D0;&#x4F9B;&#x3055;&#x308C;&#x3066;&#x8102;&#x80AA;&#x9178;&#x306B;&#x5225;&#x306E;&#x767A;&#x82BD;&#x53CD;&#x5FDC;&#x3092;&#x793A;&#x3057;&#x305F;&#x3002;&#x5909;&#x7570;&#x4F53;&#x306E;&#x767A;&#x82BD;&#x306F;&#x3001;ABA&#x3001;&#x7CD6;&#x3001;osmolites&#x3001;&#x30C8;&#x30E9;&#x30F3;&#x30B9;&#x30B8;&#x30A7;&#x30CB;&#x30C3;&#x30AF;&#x7CFB;&#x7D71;&#x306E;&#x5897;&#x52A0;WRI1&#x767A;&#x73FE;&#x306B;&#x3088;&#x3063;&#x3066;&#x7DE9;&#x548C;&#x3055;&#x308C;&#x305F;&#x52B9;&#x679C;&#x306B;&#x654F;&#x611F;&#x3067;&#x3042;&#x3063;&#x305F;&#x3002; ABA&#x5FDC;&#x7B54;&#x6027;&#x907A;&#x4F1D;&#x5B50;AtEM6&#x3068;ABA&#x3068;&#x5C0F;&#x6587;&#x5B57;&#x3092;&#x533A;&#x5225;&#x3057;&#x306A;&#x3044;3&#xFF08;ABI3&#xFF09;&#x306E;&#x767A;&#x73FE;&#x304C;wri1-1&#x5909;&#x7570;&#x4F53;&#x3067;&#x5897;&#x52A0;&#x3057;&#x305F;&#x3002; WRI1&#x3068;ABI3&#x3001;&#x7A2E;&#x5B50;&#x306E;ABA&#x5FDC;&#x7B54;&#x3092;&#x4EF2;&#x4ECB;&#x3059;&#x308B;&#x8EE2;&#x5199;&#x56E0;&#x5B50;&#x306F;&#x3001;&#x4E26;&#x5217;&#x7D4C;&#x8DEF;&#x306E;&#x884C;&#x70BA;&#x3067;&#x3042;&#x308B;&#x3053;&#x3068;&#x304C;&#x793A;&#x5506;ABI3-3&#x3068;wri1-1&#x306E;&#x9593;&#x306E;&#x4E8C;&#x91CD;&#x5909;&#x7570;&#x4F53;&#x306E;&#x89E3;&#x6790;&#x3002; 2  - &#x30C7;&#x30AA;&#x30AD;&#x30B7;&#x30B0;&#x30EB;&#x30B3;&#x30FC;&#x30B9;&#x306E;&#x6DFB;&#x52A0;&#x306F;&#x7A2E;&#x5B50;&#x767A;&#x82BD;&#x3092;&#x963B;&#x5BB3;&#x3057;&#x305F;&#x304C;&#x3001;&#x884C;&#x3092;&#x904E;&#x5270;&#x767A;&#x73FE;WRI1&#x3067;&#x305D;&#x308C;&#x4EE5;&#x4E0B;&#x3067;&#x3057;&#x305F;&#x3002;&#x82D7;&#x78BA;&#x7ACB;&#x306F;wri1-1&#x5909;&#x7570;&#x4F53;&#x3067;&#x306F;&#x6E1B;&#x5C11;&#x3057;&#x307E;&#x3057;&#x305F;&#x304C;&#x3001;&#x30B7;&#x30E7;&#x7CD6;&#x306B;&#x3088;&#x3063;&#x3066;&#x8EFD;&#x6E1B;&#x3059;&#x308B;&#x3053;&#x3068;&#x304C;&#x3067;&#x304D;&#x307E;&#x3059;&#x3002;&#x767A;&#x82BD;&#x306E;&#x53EF;&#x80FD;&#x6027;&#x30B7;&#x30B0;&#x30CA;&#x30EA;&#x30F3;&#x30B0;&#x306E;&#x5F79;&#x5272;&#x304B;&#x3089;&#x96E2;&#x308C;&#x3066;&#x3001;&#x57F9;&#x5730;&#x4E2D;&#x306E;&#x7CD6;&#x306F;wri1-1&#x82D7;&#x7ACB;&#x3061;&#x306E;&#x9593;&#x306B;&#x30D6;&#x30ED;&#x30C3;&#x30AF;&#x3084;&#x30A8;&#x30CD;&#x30EB;&#x30AE;&#x30FC;&#x4F9B;&#x7D66;&#x3092;&#x69CB;&#x7BC9;&#x3059;&#x308B;&#x3088;&#x3046;&#x8981;&#x6C42;&#x3055;&#x308C;&#x305F;&#x3002;

WRI1&#x306F;&#x3001;&#x7A2E;&#x5B50;&#x306E;&#x767A;&#x82BD;&#x3068;&#x82D7;&#x7ACB;&#x3061;&#x306B;&#x5FC5;&#x8981;&#x3067;&#x3042;&#x308B;

&#xC885;&#xC790; &#xAC1C;&#xBC1C; &#xD558;&#xB294; &#xB3D9;&#xC548; &#xC800;&#xC7A5;&#xC18C; &#xBCF5;&#xD569; &#xCD95;&#xC801; &#xC0DD;&#xC874;&#xC744; &#xC704;&#xD55C; &#xC2DD;&#xBB3C;&#xC758; &#xB2E4;&#xC74C; &#xC138;&#xB300;&#xB97C; &#xC900;&#xBE44;&#xD569;&#xB2C8;&#xB2E4;. &#xD504;&#xB85C;&#xC138;&#xC2A4; &#xADDC;&#xC815; &#xBC0F; &#xC885;&#xC790; &#xAC1C;&#xBC1C; &#xACF0; &#xAD00;&#xB828;&#xC131; &#xBC1C; &#xC544; &#xBC0F; &#xC885;&#xBB18; &#xC124;&#xB9BD;&#xC744; &#xD558;&#xB294; &#xB3D9;&#xC548; &#xC2A4;&#xD1A0;&#xB9AC;&#xC9C0; &#xBCF5;&#xD569; &#xCD95;&#xC801;&#xC758; &#xD569;&#xC131;&#xC5D0; &#xAD00;&#xC5EC; &#xD558;&#xB294; &#xB530;&#xB77C;&#xC11C;. Wrinkled1 (wri1) &#xB3CC;&#xC5F0;&#xBCC0;&#xC774; Arabidopsis (Arabidopsis thaliana)&#xC758; &#xC528;&#xC557; &#xC624;&#xC77C; &#xCD95;&#xC801;&#xC5D0; &#xC7A5;&#xC560;. WRI1 &#xC720;&#xC804;&#xC790; &#xAC1C;&#xBC1C; &#xC528;&#xC557;&#xC5D0; &#xD2B9;&#xD788; &#xD574;&#xB2F9; &#xBD84;&#xD574; &#xB300;&#xC0AC;&#xC758; &#xC81C;&#xC5B4;&#xC5D0; &#xAD00;&#xC5EC; &#xD558;&#xB294; APETALA2/&#xC5D0;&#xD2F8;&#xB80C; &#xBC18;&#xC751; &#xC694;&#xC18C; &#xBC14;&#xC778;&#xB529; &#xB2E8;&#xBC31;&#xC9C8; &#xC804;&#xC0AC; &#xC694;&#xC18C;&#xB97C; &#xC778;&#xCF54;&#xB529;&#xD569;&#xB2C8;&#xB2E4;. &#xC5EC;&#xAE30; &#xC6B0;&#xB9AC;&#xAC00; &#xBE44;&#xAD50; wri1 1 &#xB3CC;&#xC5F0;&#xBCC0;&#xC774; &#xC720;&#xC804;&#xC790; &#xBCC0;&#xD615; &#xB77C;&#xC778; wri1 1 &#xB3CC;&#xC5F0;&#xBCC0;&#xC774; &#xBC30;&#xACBD; &#xBC0F; &#xC57C;&#xC0DD; &#xD0C0;&#xC785; WRI1 &#xC57C;&#xC0DD;-&#xD0C0;&#xC785; Cdna&#xB97C; &#xD45C;&#xD604; &#xD558; &#xC5EC; &#xC885;&#xC790; &#xBC1C; &#xC544; &#xBC0F; &#xC885;&#xBB18; &#xC124;&#xB9BD;&#xC5D0;&#xC774; &#xADDC;&#xC81C; &#xC694;&#xC778;&#xC758; &#xC5ED;&#xD560;&#xC744; &#xC870;&#xC0AC; &#xD569;&#xB2C8;&#xB2E4;. &#xC2DD;&#xBB3C; WRI1 &#xC720;&#xC804;&#xC790;&#xC758; &#xD45C;&#xD604;&#xC5D0; &#xBCC0;&#xACBD; &#xB2E4;&#xB978; &#xBC1C; &#xC544; &#xC751;&#xB2F5; &#xC99D;&#xAC00;&#xC728; abscisic &#xC0B0; (ABA), &#xC124;&#xD0D5;, &#xBC0F; &#xB9E4;&#xCCB4;&#xC5D0;&#xC11C; &#xC81C;&#xACF5; &#xD558;&#xB294; &#xC9C0;&#xBC29;&#xC0B0;&#xC744; &#xBCF4;&#xC600;&#xB2E4;. &#xBC1C; &#xC544;&#xC758; &#xB3CC;&#xC5F0;&#xBCC0;&#xC774; ABA, &#xC124;&#xD0D5;, &#xBC0F; osmolites, &#xC720;&#xC804;&#xC790; &#xBCC0;&#xD615; &#xB77C;&#xC778;&#xC5D0; &#xC788;&#xB294; &#xC99D;&#xAC00; WRI1 &#xC2DD;&#xC73C;&#xB85C; &#xC644;&#xD654; &#xD558;&#xB294; &#xD6A8;&#xACFC;&#xC5D0; &#xB354; &#xBBFC;&#xAC10;&#xD55C;. ABA &#xBC18;&#xC751; &#xC720;&#xC804;&#xC790; AtEM6 &#xBC0F; ABA &#xAD6C;&#xBD84; 3 &#xC2DD; (ABI3) wri1-1 &#xB3CC;&#xC5F0;&#xBCC0;&#xC774;&#xC5D0; &#xC99D;&#xAC00; &#xD588;&#xB2E4;. Abi3-3 wri1-1 &#xC0AC;&#xC774;&#xC758; &#xC774;&#xC911; &#xB3CC;&#xC5F0;&#xBCC0;&#xC774; &#xBD84;&#xC11D; &#xC81C;&#xC548; WRI1 &#xBC0F; ABI3, &#xC885;&#xC790;&#xC758; ABA &#xC751;&#xB2F5; &#xC911;&#xC7AC; &#xD55C; &#xC804;&#xC0AC; &#xC694;&#xC18C; &#xD589;&#xB3D9;&#xC5D0; &#xBCD1;&#xB82C; &#xACBD;&#xB85C;. 2 Deoxyglucose&#xC758; &#xC885;&#xC790; &#xBC1C; &#xC544; &#xC800;&#xD574; &#xD588;&#xC9C0;&#xB9CC; &#xB108;&#xBB34; &#xB35C; &#xB77C;&#xC778; WRI1 overexpressing. &#xC885;&#xBB18; &#xC124;&#xB9BD; wri1 1 &#xB3CC;&#xC5F0;&#xBCC0;&#xC774;&#xC5D0; &#xAC10;&#xC18C; &#xD588;&#xB2E4; &#xD558;&#xC9C0;&#xB9CC; &#xC790;&#xB2F9;&#xC5D0; &#xC758;&#xD574; &#xC644;&#xD654; &#xB420; &#xC218; &#xC788;&#xC2B5;&#xB2C8;&#xB2E4;. &#xBC1C; &#xC544;&#xC5D0;&#xB294; &#xAC00;&#xB2A5;&#xD55C; &#xC2E0;&#xD638; &#xC5ED;&#xD560; &#xC678;&#xC5D0;&#xB3C4; &#xC911;&#xAC04;&#xC5D0; &#xC124;&#xD0D5; &#xBE4C;&#xB529; &#xBE14;&#xB85D; &#xBC0F; &#xC5D0;&#xB108;&#xC9C0; &#xACF5;&#xAE09; &#xC911; wri1-1 &#xBAA8; &#xC885; &#xC124;&#xB9BD; &#xD544;&#xC694; &#xD588;&#xB2E4;.

Wri1&#xB294; &#xC885;&#xC790; &#xBC1C; &#xC544; &#xBC0F; &#xC885;&#xBB18; &#xC124;&#xB9BD; &#xD544;&#xC694; &#xD569;&#xB2C8;&#xB2E4;.

La acumulaci&#xF3;n de almacenamiento compuesto durante el desarrollo de semillas se prepara la pr&#xF3;xima generaci&#xF3;n de plantas para sobrevivir. Por lo tanto, los procesos implicados en la regulaci&#xF3;n y la s&#xED;ntesis de la acumulaci&#xF3;n de almacenamiento compuesto de relevancia durante el desarrollo de las semillas de oso para la germinaci&#xF3;n y el establecimiento de las pl&#xE1;ntulas. El wrinkled1 (wri1) mutante de Arabidopsis (Arabidopsis thaliana) se ve afectada en la acumulaci&#xF3;n de aceite de semilla. El gen codifica una WRI1 APETALA2/ethylene-responsive elemento de uni&#xF3;n a la prote&#xED;na factor de transcripci&#xF3;n implicado en el control del metabolismo, particularmente la glic&#xF3;lisis, en las semillas en desarrollo. Aqu&#xED; se investiga el papel de este factor regulador en la germinaci&#xF3;n de semillas y establecimiento de pl&#xE1;ntulas mediante la comparaci&#xF3;n de los wri1-1 mutantes, las l&#xED;neas transg&#xE9;nicas que expresan el WRI1 cDNA de tipo salvaje en el fondo wri1-1 mutante y el tipo salvaje. Las plantas alteradas en la expresi&#xF3;n del gen WRI1 mostraron diferentes respuestas de germinaci&#xF3;n en el factor de crecimiento &#xE1;cido absc&#xED;sico (ABA), az&#xFA;cares, y &#xE1;cidos grasos previstas en el medio. La germinaci&#xF3;n de los mutantes fue m&#xE1;s sensible a la ABA, az&#xFA;cares y osmolites, un efecto que fue aliviada por el aumento de WRI1 expresi&#xF3;n en las l&#xED;neas transg&#xE9;nicas. La expresi&#xF3;n de genes de respuesta a ABA AtEM6 y ABA insensible a 3 (ABI3) se increment&#xF3; en el mutante wri1-1. Doble mutante an&#xE1;lisis entre ABI3-3 y wri1 1-sugiri&#xF3; que WRI1 y ABI3, un factor de transcripci&#xF3;n mediar respuestas ABA en las semillas, act&#xFA;an en las v&#xED;as paralelas. La adici&#xF3;n de 2-desoxiglucosa inhibe la germinaci&#xF3;n de semillas, pero lo hizo menos en las l&#xED;neas de sobreexpresi&#xF3;n de WRI1. Establecimiento de las pl&#xE1;ntulas se redujo en el mutante wri1-1, pero podr&#xED;a ser aliviado por la sacarosa. Adem&#xE1;s de una funci&#xF3;n de se&#xF1;alizaci&#xF3;n posible en la germinaci&#xF3;n, los az&#xFA;cares en el medio se requiere como bloques de construcci&#xF3;n y suministro de energ&#xED;a durante wri1-1 pl&#xE1;ntulas establecimiento.

WRI1 se requiere para la germinaci&#xF3;n de semillas y establecimiento de pl&#xE1;ntulas

Human U6 small nuclear RNA gene transcription by RNA polymerase III requires the general transcription factor SNAP(C), which binds to human small nuclear RNA core promoter elements and nucleates pre-initiation complex assembly with the Brf2-TFIIIB complex. Multiple components in this pathway are phosphorylated by the protein kinase CK2, including the Bdp1 subunit of the Brf2-TFIIIB complex, and RNA polymerase III, with negative and positive outcomes for U6 transcription, respectively. However, a role for CK2 phosphorylation of SNAP(C) in U6 transcription has not been defined. In this report, we investigated the role of CK2 in modulating the transcriptional properties of SNAP(C) and demonstrate that within SNAP(C), CK2 phosphorylates the N-terminal half of the SNAP190 subunit at two regions (amino acids 20-63 and 514-545) that each contain multiple CK2 consensus sites. SNAP190 phosphorylation by CK2 inhibits both SNAP(C) DNA binding and U6 transcription activity. Mutational analyses of SNAP190 support a model wherein CK2 phosphorylation triggers an allosteric inhibition of the SNAP190 Myb DNA binding domain.

The protein kinase CK2 phosphorylates SNAP190 to negatively regulate SNAPC DNA binding and human U6 transcription by RNA polymerase III.

C(4) plants have up to 10-fold higher apparent CO(2) assimilation rates than the most productive C(3) plants. This requires higher fluxes of metabolic intermediates across the chloroplast envelope membranes of C(4) plants in comparison with those of C(3) plants. In particular, the fluxes of metabolites involved in the biochemical inorganic carbon pump of C(4) plants, such as malate, pyruvate, oxaloacetate, and phosphoenolpyruvate, must be considerably higher in C(4) plants because they exceed the apparent rate of photosynthetic CO(2) assimilation, whereas they represent relatively minor fluxes in C(3) plants. While the enzymatic steps involved in the C(4) biochemical inorganic carbon pump have been studied in much detail, little is known about the metabolite transporters in the envelope membranes of C(4) chloroplasts. In this study, we used comparative proteomics of chloroplast envelope membranes from the C(3) plant pea (Pisum sativum) and mesophyll cell chloroplast envelopes from the C(4) plant maize (Zea mays) to analyze the adaptation of the mesophyll cell chloroplast envelope proteome to the requirements of C(4) photosynthesis. We show that C(3)- and C(4)-type chloroplasts have qualitatively similar but quantitatively very different chloroplast envelope membrane proteomes. In particular, translocators involved in the transport of triosephosphate and phosphoenolpyruvate as well as two outer envelope porins are much more abundant in C(4) plants. Several putative transport proteins have been identified that are highly abundant in C(4) plants but relatively minor in C(3) envelopes. These represent prime candidates for the transport of C(4) photosynthetic intermediates, such as pyruvate, oxaloacetate, and malate.

Comparative proteomics of chloroplast envelopes from C3 and C4 plants reveals specific adaptations of the plastid envelope to C4 photosynthesis and candidate proteins required for maintaining C4 metabolite fluxes.

The phloem plays a crucial role in assimilate and nutrient transport, pathogen response, and plant growth and development. Yet, few species have yielded pure phloem exudate and, if proteins need to be analysed, those species may not have sequenced genomes, making identification difficult. The enrichment of Arabidopsis thaliana phloem exudate in amounts large enough to allow for metabolite and protein analysis is described. Using this method, it was possible to identify 65 proteins present in the Arabidopsis phloem exudate. The majority of these proteins could be grouped by response to pathogens, stress, or hormones, carbon metabolism, protein interaction, modification, and turnover, and transcription factors. It was also possible to detect 11 proteins that play a role in lipid/fatty acid metabolism (aspartic protease, putative 3-β-hydroxysteroid dehydrogenase, UDP-sulphoquinovose synthase/SQD1, lipase, PIG-P-like protein: phosphatidylinositol-N-acetylglucosaminyltransferase), storage (glycine-rich protein), binding (annexin, lipid-associated family protein, GRP17/oleosin), and/or signalling (annexin, putative lipase, PIG-P-like protein). Along with putative lipid-binding proteins, several lipids and fatty acids could be identified. Only a few examples exist of lipids (jasmonic acid, oxylipins) or lipid-binding proteins (DIR1, acyl-CoA-binding protein) in the phloem. Finding hydrophobic compounds in an aqueous environment is not without precedence in biological systems: human blood contains a variety of lipids, many of which play a significant role in human health. In blood, lipids are transported while bound to proteins. The present findings of lipids and lipid-binding proteins in phloem exudates suggest that a similar long-distance lipid signalling exists in plants and may play an important role in plant growth and development.

Identification of lipids and lipid-binding proteins in phloem exudates from Arabidopsis thaliana.

Plants are sessile and cannot move to appropriate hiding places or feeding grounds to escape adverse conditions. As a consequence, they evolved mechanisms to detect changes in their environment, communicate these to different organs, and adjust development accordingly. These adaptations include two long-distance transport systems which are essential in plants: the xylem and the phloem. The phloem serves as a major trafficking pathway for assimilates, viruses, RNA, plant hormones, metabolites, and proteins with functions ranging from synthesis to metabolism to signaling. The study of signaling compounds within the phloem is essential for our understanding of plant communication of environmental cues. Determining the nature of signals and the mechanisms by which they are communicated through the phloem will lead to a more complete understanding of plant development and plant responses to stress. In our analysis of Arabidopsis phloem exudates, we had identified several lipid-binding proteins as well as fatty acids and lipids. The latter are not typically expected in the aqueous environment of sieve elements. Hence, lipid transport in the phloem has been given little attention until now. Long-distance transport of hydrophobic compounds in an aqueous system is not without precedence in biological systems: a variety of lipids is found in human blood and is often bound to proteins. Some lipid-protein complexes are transported to other tissues for storage, use, modification, or degradation; others serve as messengers and modulate transcription factor activity. By simple analogy it raises the possibility that lipids and the respective lipid-binding proteins in the phloem serve similar functions in plants and play an important role in stress and developmental signaling. Here, we introduce the lipid-binding proteins and the lipids we found in the phloem and discuss the possibility that they may play an important role in developmental and stress signaling.

New aspects of Phloem-mediated long-distance lipid signaling in plants.

As the world population grows, our need for food increases drastically. Limited amounts of arable land lead to a competition between food and fuel crops, while changes in the global climate may impact future crop yields. Thus, a second "green revolution" will need a better understanding of the processes essential for plant growth and development. One approach toward the solution of this problem is to better understand regulatory and transport processes in C4 plants. C4 plants display an up to 10-fold higher apparent CO2 assimilation and higher yields while maintaining high water use efficiency. This requires differential regulation of mesophyll (M) and bundle sheath (BS) chloroplast development as well as higher metabolic fluxes of photosynthetic intermediates between cells and particularly across chloroplast envelopes. While previous analyses of overall chloroplast membranes have yielded significant insight, our comparative proteomics approach using enriched BS and M chloroplast envelopes of Zea mays allowed us to identify 37 proteins of unknown function that have not been seen in these earlier studies. We identified 280 proteins, 84% of which are known/predicted to be present in chloroplasts. Seventy-four percent have a known or predicted membrane association. Twenty-one membrane proteins were 2-15 times more abundant in BS cells, while 36 of the proteins were more abundant in M chloroplast envelopes. These proteins could represent additional candidates of proteins essential for development or metabolite transport processes in C4 plants. RT-PCR confirmed differential expression of 13 candidate genes. Chloroplast association for seven proteins was confirmed using YFP/GFP labeling. Gene expression of four putative transporters was examined throughout the leaf and during the greening of leaves. Genes for a PIC-like protein and an ER-AP-like protein show an early transient increase in gene expression during the transition to light. In addition, PIC gene expression is increased in the immature part of the leaf and was lower in the fully developed parts of the leaf, suggesting a need for/incorporation of the protein during chloroplast development.

Comparative proteomics of chloroplasts envelopes from bundle sheath and mesophyll chloroplasts reveals novel membrane proteins with a possible role in c4-related metabolite fluxes and development.

The epidermis is not only the interphase between the plant and the environment but also a growth-limiting tissue. Understanding the initiation and regulation of its expansion growth is essential for addressing the need for more food and fuel. We used mass spectrometry to identify proteins from auxin (indole-3-acetic acid; IAA)-induced rapidly growing corn (Zea mays) coleoptiles to find possible candidates controlling this growth as well as the underlying cell wall and cuticle biosynthesis. Excised sections were incubated for 4 h in the absence or presence of IAA, protein extracted, and analyzed using LC-ESI-MS/MS. Of 86 proteins identified, 15 showed a predicted association with cell wall/cuticle biosynthesis or trafficking machinery; four identifications revealed novel proteins of unknown function. In parallel, real-time PCR indicated that the steady-state mRNA levels of genes with a known or predicted role in cell-wall biosynthesis increase upon treatment with auxin. Importantly, genes encoding two of the hypothetical proteins also show higher levels of mRNA; additionally, their gene expression is down-regulated as coleoptile growth ceases and up-regulated in expanding leaves. This suggests a major role of those novel proteins in the regulation of processes related to cell and organ expansion and thus plant growth.

Susanne Hoffmann-Benning

Biochemistry and Molecular Biology

Michigan State Universtiy

Collection and Analysis of Arabidopsis Phloem Exudates Using the EDTA-facilitated Method

Susanne Hoffmann-Benning

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Biochemistry and Molecular Biology

Michigan State Universtiy

Collection and Analysis of Arabidopsis Phloem Exudates Using the EDTA-facilitated Method

Biochemistry and Molecular Biology