fluorescence-time-lapse-imaging-complete-s-venezuelae-life-cycle

Fluorescence Time-lapse Imaging of the Complete <em>S. venezuelae</em> Life Cycle Using a Microfluidic Device

Live-cell imaging of biological processes at the single cell level has been instrumental to our current understanding of the subcellular organization of ...

John Innes Centre, Norwich Research Park

&#x5728;&#x7EC6;&#x83CC;&#x4E2D;&#xFF0C;&#x80DE;&#x8D28;&#x5206;&#x88C2;&#x662F;&#x7EC6;&#x80DE;&#x58C1;&#x7684;&#x4F9D;&#x8D56;&#x4FBF;&#x5229;&#x5728;&#x6536;&#x7F29;&#x671F;&#x95F4;&#x6539;&#x5EFA;&#x7684;&#x6EB6;&#x89E3;&#x9176;&#x3002;&#x5728;&#x8FD9;&#x9879;&#x5DE5;&#x4F5C;&#xFF0C;&#x6211;&#x4EEC;&#x627E;&#x51FA;&#x8FC4;&#x4ECA;&#x65E0;&#x5468;&#x8D28;&#x86CB;&#x767D;&#xFF0C;&#x8005;&#xFF0C;&#x6240;&#x9700;&#x7684;&#x7EC6;&#x80DE;&#x5206;&#x88C2;&#x548C;&#x5728; Caulobacter crescentus &#x7684;&#x6781;&#x6027;&#x3002;&#x8005;&#x662F;&#x7531;&#x56DB;&#x4E2A;&#x80BD;&#x805A;&#x7CD6;&#x5177;&#x6709;&#x7EA6;&#x675F;&#x529B; &#xFF08;LysM&#xFF09; &#x57DF;&#x548C; C &#x7EC8;&#x7AEF;&#x6EB6;&#x8461;&#x8404;&#x7403;&#x83CC;&#x9176;&#x6837; &#xFF08;LytM&#xFF09; &#x80BD;&#x9176;&#x57DF;&#x7EC4;&#x6210;&#x3002;&#x5B83;&#x5C06;&#x7ED1;&#x5B9A;&#x5230;&#x5B64;&#x7ACB;&#x7684; murein &#x4E00;&#x4F53;&#x5916;&#xFF0C;&#x548C;&#x62DB;&#x8058;&#x5230;&#x901A;&#x8FC7;&#x4E0E;&#x7EC6;&#x80DE;&#x5206;&#x88C2;&#x86CB;&#x767D; FtsN.&#x7A81;&#x53D8;&#x5206;&#x6790;&#x8868;&#x660E; LysM &#x57DF;&#x662F;&#x5FC5;&#x8981;&#x548C;&#x8DB3;&#x591F;&#x7684;&#x672C;&#x5730;&#x5316;&#x8005;&#xFF0C;&#x867D;&#x7136;&#x5176;&#x80BD;&#x9176;&#x57DF;&#x662F;&#x975E;&#x5E38;&#x91CD;&#x8981;&#x7684;&#x529F;&#x80FD;&#x8FDB;&#x884C;&#x4EA4;&#x4E92;&#x7684;&#x72ED;&#x7A84;&#x7684;&#x7AD9;&#x70B9;&#x3002;&#x7B26;&#x5408;&#x5728;&#x7EC6;&#x80DE;&#x58C1;&#x6C34;&#x89E3;&#x4F5C;&#x7528;&#xFF0C;&#x8005;&#x88AB;&#x53D1;&#x73B0;&#x4E0E;&#x7EAF;&#x51C0;&#x7684; murein &#x4E00;&#x4F53;&#x5916;&#x8FDB;&#x884C;&#x4EA4;&#x4E92;&#xFF0C;&#x5E76;&#x4FC3;&#x4F7F;&#x7EC6;&#x80DE;&#x88C2;&#x89E3;&#x540E;&#x7684;&#x751F;&#x4EA7;&#x8FC7;&#x5269;&#x3002;&#x5176;&#x6740;&#x706D;&#x5BFC;&#x81F4;&#x5916;&#x819C;&#x5185;&#x9677;&#xFF0C;&#x9020;&#x6210;&#x91CD;&#x5927;&#x5EF6;&#x8BEF;&#x7EC6;&#x80DE;&#x8D28;&#x5206;&#x88C2;&#x548C;&#x6700;&#x540E;&#x5206;&#x79BB;&#x7684;&#x5B50;&#x7EC6;&#x80DE;&#x4E4B;&#x95F4;&#x4E2D;&#x7684;&#x4E25;&#x91CD;&#x7F3A;&#x9677;&#x3002;&#x603B;&#x4F53;&#x800C;&#x8A00;&#xFF0C;&#x8FD9;&#x4E9B;&#x7ED3;&#x679C;&#x8868;&#x660E;&#x8005;&#x662F;&#x5468;&#x8D28; C.crescentus divisome&#xFF0C;&#x4FBF;&#x4E8E;&#x6539;&#x5EFA;&#x7684;&#x80BD;&#x805A;&#x7CD6;&#x5C42;&#xFF0C;&#x56E0;&#x6B64;&#xFF0C;&#x534F;&#x8C03;&#x53F8;&#x8FC7;&#x7A0B;&#x4E2D;&#x7684;&#x5355;&#x5143;&#x683C;&#x4FE1;&#x5C01;&#x7F29;&#x7A84;&#x7684;&#x7EC4;&#x6210;&#x90E8;&#x5206;&#x3002;

&#x8005;&#xFF0C;&#x6240;&#x9700;&#x7684;&#x80BD;&#x805A;&#x7CD6;&#x5728; Caulobacter crescentus &#x4E2D;&#x7684;&#x7EC6;&#x80DE;&#x5206;&#x88C2;&#x8FC7;&#x7A0B;&#x91CD;&#x6784;&#x4E00;&#x4E2A;&#x65B0;&#x7684;&#x56E0;&#x7D20;&#x3002;

In bacteria, cytokinesis is dependent on lytic enzymes that facilitate remodelling of the cell wall during constriction. In this work, we identify a thus far uncharacterized periplasmic protein, DipM, that is required for cell division and polarity in Caulobacter crescentus. DipM is composed of four peptidoglycan binding (LysM) domains and a C-terminal lysostaphin-like (LytM) peptidase domain. It binds to isolated murein sacculi in vitro, and is recruited to the site of constriction through interaction with the cell division protein FtsN. Mutational analyses showed that the LysM domains are necessary and sufficient for localization of DipM, while its peptidase domain is essential for function. Consistent with a role in cell wall hydrolysis, DipM was found to interact with purified murein sacculi in vitro and to induce cell lysis upon overproduction. Its inactivation causes severe defects in outer membrane invagination, resulting in a significant delay between cytoplasmic compartmentalization and final separation of the daughter cells. Overall, these findings indicate that DipM is a periplasmic component of the C. crescentus divisome that facilitates remodelling of the peptidoglycan layer and, thus, coordinated constriction of the cell envelope during the division process.

DipM, a new factor required for peptidoglycan remodelling during cell division in Caulobacter crescentus.

Chez les bact&#xE9;ries, la cytocin&#xE8;se est d&#xE9;pendante des enzymes lytiques qui facilitent le remodelage de la paroi cellulaire au cours de la constriction. Dans ce travail, nous identifions une prot&#xE9;ine jusqu&#x26;#39;ici non caract&#xE9;ris&#xE9;e p&#xE9;riplasmique, DIPM, ce qui est n&#xE9;cessaire pour la division cellulaire et de la polarit&#xE9; dans Caulobacter crescentus. DIPM est compos&#xE9; de quatre peptidoglycane de liaison (LysM) domaines et une extr&#xE9;mit&#xE9; C-terminale du domaine en forme de lysostaphine peptidase (LytM). Il se lie &#xE0; murein isol&#xE9; sacculi in vitro, et est recrut&#xE9; pour le site de constriction &#xE0; travers l&#x26;#39;interaction avec le FtsN prot&#xE9;ines de division cellulaire. Analyses mutationnelles ont montr&#xE9; que les domaines LysM sont n&#xE9;cessaires et suffisants pour la localisation de DIPM, tandis que son domaine peptidase est essentiel pour la fonction. Conform&#xE9;ment &#xE0; un r&#xF4;le dans l&#x26;#39;hydrolyse paroi cellulaire, DIPM a &#xE9;t&#xE9; trouv&#xE9; pour interagir avec purifi&#xE9; murein sacculi in vitro et d&#x26;#39;induire la lyse des cellules sur la surproduction. Son inactivation provoque de graves d&#xE9;fauts dans la membrane externe invagination, ce qui entra&#xEE;ne un retard important entre le cloisonnement cytoplasmique et la s&#xE9;paration finale des cellules filles. Dans l&#x26;#39;ensemble, ces r&#xE9;sultats indiquent que DIPM est un composant p&#xE9;riplasmique de la divisome C. crescentus qui facilite le remodelage de la couche de peptidoglycane et, par cons&#xE9;quent, la constriction coordonn&#xE9;e de l&#x26;#39;enveloppe cellulaire au cours du processus de division.

DIPM, un nouveau facteur requis pour peptidoglycane Remodelling Durant la division cellulaire dans Caulobacter crescentus

In Bakterien h&auml;ngt der Zellteilung lytischer Enzyme, die Umgestaltung der Zellwand w&auml;hrend Verengung erleichtern. In dieser Arbeit identifizieren wir ein bisher uncharakterisiertes periplasmatischen Protein, DipM, das f&uuml;r die Zellteilung und Polarit&auml;t in Caulobacter Crescentus erforderlich ist. DipM besteht aus vier Dom&auml;nen der Peptidoglycan-Bindung (LysM) und einer C-terminalen Lysostaphin-wie (LytM) Peptidase-Dom&auml;ne. Es bindet an isolierten Murein Sacculi in-vitro-, und auf der Website der Verengung durch Interaktion mit der Zellteilung-Protein FtsN. teilhabende Analysen zeigten, dass die LysM-Dom&auml;nen sind notwendig und hinreichend f&uuml;r die Lokalisierung der DipM, w&auml;hrend seiner Peptidase-Dom&auml;ne notwendig f&uuml;r die Funktion ist eingestellt ist. In &Uuml;bereinstimmung mit eine Rolle bei der Zellwand Hydrolyse, wurde DipM Interaktion mit gereinigtem Murein Sacculi in-vitro- und Zellenzerst&ouml;rung auf &Uuml;berproduktion veranlassen gefunden. Seine Inaktivierung f&uuml;hrt zu schwere defekten Au&szlig;enmembran Invagination, was zu einer erheblichen Verz&ouml;gerung zwischen zytoplasmatischen Untergliederung und endg&uuml;ltige Trennung von der Tochterzellen. Insgesamt zeigen diese Ergebnisse, dass DipM periplasmatischen Komponente der C. Crescentus Divisome ist, die Umgestaltung der Peptidoglycan Schicht erleichtert und somit Verengung der die Zellh&uuml;llen bei der Abteilung koordiniert.

DipM, ein neuer Faktor f&uuml;r Peptidoglycan Umbau w&auml;hrend der Zellteilung in Caulobacter Crescentus erforderlich.

Nei batteri, Citocinesi in &egrave; dipendente da enzimi litici che facilitano il rimodellamento della parete cellulare durante la costrizione. In questo lavoro, ci identifichiamo una proteina periplasmica finora atipiche, DipM, che &egrave; necessaria per la divisione cellulare e di polarit&agrave; in Caulobacter ha. DipM &egrave; composta da quattro domini di associazione peptidoglicano (LysM) e un dominio C-terminale simile a lysostaphin (LytM) peptidasi. Si lega a sacculi murein isolato in vitro e viene reclutato per il sito di costrizione attraverso l&#39;interazione con la proteina di divisione cellulare FtsN. analisi mutazionale ha dimostrato che i domini LysM sono necessarie e sufficienti per la localizzazione di DipM, mentre il suo dominio peptidasi &egrave; essenziale per la funzione. Coerente con un ruolo nella parete cellulare idrolisi, DipM &egrave; stato trovato a interagire con sacculi murein purificato in vitro e per indurre la lisi delle cellule alla sovrapproduzione. Sua inattivazione provoca gravi difetti nell&#39;invaginazione della membrana esterna, causando un notevole ritardo tra compartimentazione citoplasmatici e definitiva separazione delle cellule figlie. Nel complesso, questi risultati indicano che DipM &egrave; un componente periplasmico il c. ha divisome che facilita il rimodellamento dello strato di peptidoglicano e, quindi, coordinato costrizione della busta delle cellule durante il processo di divisione.

DipM, un nuovo fattore richiesto per peptidoglicano rimodellamento durante la divisione cellulare in Caulobacter ha.

&#x7D30;&#x83CC;&#x3067;&#x306F;&#x3001;&#x7D30;&#x80DE;&#x8CEA;&#x5206;&#x88C2;&#x306F;&#x3001;&#x53CE;&#x7E2E;&#x6642;&#x306E;&#x7D30;&#x80DE;&#x58C1;&#x306E;&#x6539;&#x9020;&#x3092;&#x5BB9;&#x6613;&#x6EB6;&#x83CC;&#x9175;&#x7D20;&#x306B;&#x4F9D;&#x5B58;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x672C;&#x7814;&#x7A76;&#x3067;&#x306F;&#x3001;&#x30AB;&#x30A6;&#x30ED;&#x30D0;&#x30AF;&#x30BF;&#x30FC;crescentus&#x306E;&#x7D30;&#x80DE;&#x5206;&#x88C2;&#x3068;&#x6975;&#x6027;&#x306E;&#x305F;&#x3081;&#x306B;&#x5FC5;&#x8981;&#x3068;&#x3055;&#x308C;&#x308B;&#x3053;&#x308C;&#x307E;&#x3067;&#x672A;&#x77E5;&#x30DA;&#x30EA;&#x30D7;&#x30E9;&#x30BA;&#x30E0;&#x86CB;&#x767D;&#x8CEA;&#x3001;DIPM&#x3092;&#x8B58;&#x5225;&#x3057;&#x307E;&#x3059;&#x3002; DIPM&#x306F;&#x3001;4&#x3064;&#x306E;&#x30DA;&#x30D7;&#x30C1;&#x30C9;&#x30B0;&#x30EA;&#x30AB;&#x30F3;&#x7D50;&#x5408;&#xFF08;LysM&#xFF09;&#x30C9;&#x30E1;&#x30A4;&#x30F3;&#x3068;C&#x672B;&#x7AEF;&#x30EA;&#x30BD;&#x30B9;&#x30BF;&#x30D5;&#x30A3;&#x30F3;&#x306E;&#x3088;&#x3046;&#x306A;&#xFF08;LytM&#xFF09;&#x30DA;&#x30D7;&#x30C1;&#x30C0;&#x30FC;&#x30BC;&#x306E;&#x30C9;&#x30E1;&#x30A4;&#x30F3;&#x3067;&#x69CB;&#x6210;&#x3055;&#x308C;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x305D;&#x308C;&#x306F;&#x3001;in vitro&#x3067;&#x5B64;&#x7ACB;&#x30E0;&#x30EC;&#x30A4;&#x30F3;&#x7403;&#x5F62;&#x56A2;&#x306B;&#x7D50;&#x5408;&#x3057;&#x3001;&#x7D30;&#x80DE;&#x5206;&#x88C2;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;FtsN&#x3068;&#x306E;&#x76F8;&#x4E92;&#x4F5C;&#x7528;&#x3092;&#x4ECB;&#x3057;&#x3066;&#x53CE;&#x7E2E;&#x306E;&#x30B5;&#x30A4;&#x30C8;&#x306B;&#x52DF;&#x96C6;&#x3057;&#x3066;&#x304A;&#x308A;&#x307E;&#x3059;&#x3002;&#x5909;&#x7570;&#x89E3;&#x6790;&#x3067;&#x306F;&#x3001;&#x305D;&#x306E;&#x30DA;&#x30D7;&#x30C1;&#x30C0;&#x30FC;&#x30BC;&#x30C9;&#x30E1;&#x30A4;&#x30F3;&#x304C;&#x6A5F;&#x80FD;&#x306B;&#x5FC5;&#x9808;&#x3067;&#x3042;&#x308B;&#x4E00;&#x65B9;LysM&#x30C9;&#x30E1;&#x30A4;&#x30F3;&#x306F;&#x3001;DIPM&#x306E;&#x30ED;&#x30FC;&#x30AB;&#x30E9;&#x30A4;&#x30BA;&#x306E;&#x305F;&#x3081;&#x306B;&#x5FC5;&#x8981;&#x304B;&#x3064;&#x5341;&#x5206;&#x3067;&#x3042;&#x308B;&#x3053;&#x3068;&#x3092;&#x793A;&#x3057;&#x305F;&#x3002;&#x7D30;&#x80DE;&#x58C1;&#x306E;&#x52A0;&#x6C34;&#x5206;&#x89E3;&#x306B;&#x304A;&#x3051;&#x308B;&#x5F79;&#x5272;&#x3068;&#x4E00;&#x81F4;&#x3057;&#x3001;DIPM&#x306F;&#x3001;in vitro&#x3067;&#x7CBE;&#x88FD;&#x30E0;&#x30EC;&#x30A4;&#x30F3;&#x7403;&#x5F62;&#x56A2;&#x3068;&#x5BFE;&#x8A71;&#x3059;&#x308B;&#x3068;&#x904E;&#x5270;&#x306B;&#x7D30;&#x80DE;&#x6EB6;&#x89E3;&#x3092;&#x8A98;&#x5C0E;&#x3059;&#x308B;&#x3053;&#x3068;&#x304C;&#x767A;&#x898B;&#x3055;&#x308C;&#x307E;&#x3057;&#x305F;&#x3002;&#x305D;&#x306E;&#x4E0D;&#x6D3B;&#x6027;&#x5316;&#x306F;&#x7D30;&#x80DE;&#x8CEA;&#x306E;&#x533A;&#x753B;&#x3068;&#x5A18;&#x7D30;&#x80DE;&#x306E;&#x6700;&#x7D42;&#x7684;&#x306A;&#x5206;&#x96E2;&#x306E;&#x9593;&#x306B;&#x6709;&#x610F;&#x306A;&#x9045;&#x5EF6;&#x304C;&#x751F;&#x3058;&#x3001;&#x5916;&#x819C;&#x306E;&#x9665;&#x5165;&#x306B;&#x304A;&#x3051;&#x308B;&#x91CD;&#x5927;&#x306A;&#x6B20;&#x9665;&#x304C;&#x767A;&#x751F;&#x3057;&#x307E;&#x3059;&#x3002;&#x5168;&#x4F53;&#x3068;&#x3057;&#x3066;&#x3001;&#x3053;&#x308C;&#x3089;&#x306E;&#x77E5;&#x898B;&#x306F;&#x3001;DIPM&#x306F;&#x9664;&#x7B97;&#x51E6;&#x7406;&#x4E2D;&#x306B;&#x30DA;&#x30D7;&#x30C1;&#x30C9;&#x30B0;&#x30EA;&#x30AB;&#x30F3;&#x5C64;&#x306E;&#x6539;&#x9020;&#x3068;&#x3001;&#x305D;&#x306E;&#x7D50;&#x679C;&#x3001;&#x7D30;&#x80DE;&#x30A8;&#x30F3;&#x30D9;&#x30ED;&#x30FC;&#x30D7;&#x306E;&#x5354;&#x8ABF;&#x53CE;&#x7E2E;&#x3092;&#x4FC3;&#x9032;&#x3059;&#x308B;C. crescentus&#x306E;divisome&#x306E;&#x30DA;&#x30EA;&#x30D7;&#x30E9;&#x30BA;&#x30E0;&#x6210;&#x5206;&#x3067;&#x3042;&#x308B;&#x3053;&#x3068;&#x3092;&#x793A;&#x3057;&#x3066;&#x3044;&#x308B;&#x3002;

DIPM&#x3001;&#x30AB;&#x30A6;&#x30ED;&#x30D0;&#x30AF;&#x30BF;&#x30FC;Crescentus&#x3067;&#x3001;&#x7D30;&#x80DE;&#x5206;&#x88C2;&#x306E;&#x9593;&#x306B;&#x30DA;&#x30D7;&#x30C1;&#x30C9;&#x30B0;&#x30EA;&#x30AB;&#x30F3;&#x30EA;&#x30E2;&#x30C7;&#x30EA;&#x30F3;&#x30B0;&#x306B;&#x5FC5;&#x8981;&#x306A;&#x65B0;&#x3057;&#x3044;&#x30D5;&#x30A1;&#x30AF;&#x30BF;&#x30FC;

&#xBC15;&#xD14C;&#xB9AC;&#xC544;, cytokinesis &#xC138;&#xD3EC; &#xBCBD;&#xC758; &#xC218;&#xCD95; &#xD558;&#xB294; &#xB3D9;&#xC548; &#xB9AC; &#xBAA8;&#xB378;&#xB9C1;&#xC744; &#xCD09;&#xC9C4; &#xD558;&#xB294; &#xC6A9;&#xADE0;&#xC131; &#xD6A8;&#xC18C;&#xC5D0; &#xC885;&#xC18D; &#xB429;&#xB2C8;&#xB2E4;. &#xC774; &#xC791;&#xD488;&#xC5D0;&#xC11C; &#xC6B0;&#xB9AC;&#xB294; &#xC9C0;&#xAE08;&#xAE4C;&#xC9C0; &#xC0C8;&#xB86D;&#xAC70;&#xB098; periplasmic &#xB2E8;&#xBC31;&#xC9C8;, &#xC138;&#xD3EC; &#xBD84;&#xC5F4; &#xBC0F; Caulobacter crescentus&#xC5D0; &#xADF9;&#xC131;&#xC5D0; &#xD544;&#xC694;&#xD55C; DipM &#xC2DD;&#xBCC4;. DipM 4 peptidoglycan &#xBC14;&#xC778;&#xB529; (LysM) &#xB3C4;&#xBA54;&#xC778; &#xBC0F; C &#xD130;&#xBBF8;&#xB110; lysostaphin &#xAC19;&#xC740; (LytM) peptidase &#xB3C4;&#xBA54;&#xC778;&#xC73C;&#xB85C; &#xAD6C;&#xC131; &#xB429;&#xB2C8;&#xB2E4;. &#xADF8;&#xAC83;&#xC740; &#xACA9;&#xB9AC; murein sacculi &#xCCB4; &#xC678;&#xC5D0; &#xBC14;&#xC778;&#xB529;&#xD558;&#xACE0; &#xC218;&#xCD95; &#xC138;&#xD3EC; &#xB2E8;&#xBC31;&#xC9C8; FtsN. Mutational &#xBD84;&#xC11D; &#xBCF4;&#xC600;&#xB2E4; peptidase &#xB3C4;&#xBA54;&#xC778; &#xAE30;&#xB2A5;&#xC744; &#xC704;&#xD574; &#xD544;&#xC218;&#xC801;&#xC785;&#xB2C8;&#xB2E4; &#xD558;&#xB294; &#xB3D9;&#xC548; LysM &#xB3C4;&#xBA54;&#xC778;&#xC740; &#xD544;&#xC694; &#xD558; &#xACE0; DipM, &#xC9C0;&#xC5ED;&#xD654;&#xC5D0; &#xB300; &#xD55C; &#xCDA9;&#xBD84; &#xD55C; &#xADF8;&#xC640; &#xC0C1;&#xD638; &#xC791;&#xC6A9;&#xC744; &#xD1B5;&#xD574; &#xC0AC;&#xC774;&#xD2B8;&#xC5D0; &#xCC44;&#xC6A9;. &#xC138;&#xD3EC; &#xBCBD; &#xAC00;&#xC218;&#xBD84;&#xD574;&#xC5D0; &#xC5ED;&#xD560;&#xC640; &#xC77C;&#xCE58;, DipM &#xBC1C;&#xACAC; &#xC815;&#xD654; murein sacculi &#xCCB4; &#xC678;&#xC640; &#xC0C1;&#xD638; &#xC791;&#xC6A9; &#xD558; &#xACE0; &#xACFC;&#xC789; &#xC2DC; &#xC138;&#xD3EC; &#xC138;&#xD3EC;&#xC758; &#xC6A9; &#xD574;&#xB97C; &#xC720;&#xBC1C; &#xD558;&#xB294; &#xAC83;. &#xBE44;&#xD65C;&#xC131;&#xD654; &#xC678;&#xBD80; &#xB9C9; invagination, &#xC138;&#xD3EC;&#xC9C8; &#xBCF4;&#xD5D8;&#xD68C;&#xC0AC;&#xC640; &#xB538; &#xC138;&#xD3EC;&#xC758; &#xCD5C;&#xC885; &#xBD84;&#xB9AC; &#xC0AC;&#xC774;&#xC758; &#xC911;&#xC694; &#xD55C; &#xC9C0;&#xC5F0;&#xC758; &#xACB0;&#xACFC;&#xB85C;&#xC758; &#xC2EC;&#xAC01;&#xD55C; &#xACB0;&#xD568; &#xBC1C;&#xC0DD; &#xD569;&#xB2C8;&#xB2E4;. &#xC804;&#xBC18;&#xC801;&#xC73C;&#xB85C;, &#xC774;&#xB7EC;&#xD55C; &#xACB0;&#xACFC; DipM peptidoglycan &#xACC4;&#xCE35;&#xC758; &#xB9AC; &#xBAA8;&#xB378;&#xB9C1;&#xC744; &#xC6A9;&#xC774; &#xD558; &#xAC8C; &#xD558; &#xACE0; &#xB530;&#xB77C;&#xC11C; &#xC870;&#xC815; &#xBD80;&#xBB38; &#xACFC;&#xC815; &#xC140; &#xBD09;&#xD22C;&#xC758; &#xC218;&#xCD95; &#xD558;&#xB294; C. crescentus divisome&#xC758; periplasmic &#xAD6C;&#xC131; &#xC694;&#xC18C; &#xC784;&#xC744; &#xB098;&#xD0C0;&#xB0C5;&#xB2C8;&#xB2E4;.

DipM peptidoglycan &#xAC1C;&#xC7A5; Caulobacter crescentus&#xC5D0;&#xC11C; &#xC138;&#xD3EC; &#xBD84;&#xC5F4; &#xB3D9;&#xC548;&#xC5D0; &#xD544;&#xC694;&#xD55C; &#xC0C8;&#xB85C;&#xC6B4; &#xC694;&#xC18C;.

En las bacterias, citocinesis es dependiente de las enzimas l&#xED;ticas que faciliten la remodelaci&#xF3;n de la pared celular durante la constricci&#xF3;n. En este trabajo, se identifican una prote&#xED;na hasta ahora no caracterizada peripl&#xE1;smico, DIPM, lo que se requiere para la divisi&#xF3;n celular y la polaridad en Caulobacter crescentus. DIPM se compone de cuatro peptidoglicano vinculantes (LysM) dominios y un C-terminal-como lysostaphin (LytM) de dominio peptidasa. Se une a sacculi mure&#xED;na aislado in vitro, y ha sido contratado para el sitio de la constricci&#xF3;n trav&#xE9;s de la interacci&#xF3;n con la prote&#xED;na de la c&#xE9;lula FtsN divisi&#xF3;n. An&#xE1;lisis mutacional mostr&#xF3; que los dominios LysM son necesarias y suficientes para la localizaci&#xF3;n de DIPM, mientras que su dominio peptidasa es esencial para la funci&#xF3;n. De acuerdo con un papel en la hidr&#xF3;lisis de la pared celular, DIPM se encontr&#xF3; a interactuar con la mure&#xED;na purificada sacculi in vitro e inducir la lisis celular a la sobreproducci&#xF3;n. Su inactivaci&#xF3;n causa defectos graves en invaginaci&#xF3;n membrana externa, resultando en un retraso significativo entre compartimentaci&#xF3;n citopl&#xE1;smica y separaci&#xF3;n final de las c&#xE9;lulas hijas. En general, estos resultados indican que DIPM es un componente peripl&#xE1;smico de la divisome C. crescentus que facilita la remodelaci&#xF3;n de la capa de peptidoglicano y, por tanto, la constricci&#xF3;n coordinada de la envoltura celular durante el proceso de divisi&#xF3;n.

DIPM, un nuevo factor necesario para el peptidoglicano Remodelaci&#xF3;n Durante la divisi&#xF3;n celular en la Caulobacter crescentus

In eukaryotes, the differentiation of cellular extensions such as cilia or neuronal axons depends on the partitioning of proteins to distinct plasma membrane domains by specialized diffusion barriers. However, examples of this compartmentalization strategy are still missing for prokaryotes, although complex cellular architectures are also widespread among this group of organisms. This study reveals the existence of a protein-mediated membrane diffusion barrier in the stalked bacterium Caulobacter crescentus. We show that the Caulobacter cell envelope is compartmentalized by macromolecular complexes that prevent the exchange of both membrane and soluble proteins between the polar stalk extension and the cell body. The barrier structures span the cross-sectional area of the stalk and comprise at least four proteins that assemble in a cell-cycle-dependent manner. Their presence is critical for cellular fitness because they minimize the effective cell volume, allowing faster adaptation to environmental changes that require de novo synthesis of envelope proteins.

General protein diffusion barriers create compartments within bacterial cells.

Bacterial cell division involves the dynamic assembly of division proteins and coordinated constriction of the cell envelope. A wide range of factors regulates cell division--including growth and environmental stresses--and the targeting of the division machinery has been a widely discussed approach for antimicrobial therapies. This paper introduces divin, a small molecule inhibitor of bacterial cell division that may facilitate mechanistic studies of this process. Divin disrupts the assembly of late division proteins, reduces peptidoglycan remodeling at the division site, and blocks compartmentalization of the cytoplasm. In contrast to other division inhibitors, divin does not interact with the tubulin homologue FtsZ, affect chromosome segregation, or activate regulatory mechanisms that inhibit cell division indirectly. Our studies of bacterial cell division using divin as a probe suggest that dividing bacteria proceed through several morphological stages of the cell envelope, and FtsZ is required but not sufficient to compartmentalize the cytoplasmic membrane at the division site. Divin is only moderately toxic to mammalian cells at concentrations that inhibit the growth of clinical pathogens. These characteristics make divin a useful probe for studying bacterial cell division and a starting point for the development of new classes of therapeutic agents.

Divin: a small molecule inhibitor of bacterial divisome assembly.

The Gram-negative bacterium Caulobacter crescentus forms a thin polar stalk, which mediates its attachment to solid surfaces. Whereas stalks remain short (1 µm) in nutrient-rich conditions, they lengthen dramatically (up to 30 µm) upon phosphate starvation. A long-standing hypothesis is that the Caulobacter stalk functions as a nutrient scavenging "antenna" that facilitates phosphate uptake and transport to the cell body. The mechanistic details of this model must be revisited, given our recent identification of a protein-mediated diffusion barrier, which prevents the exchange of both membrane and soluble proteins between the stalk extension and the cell body. In this report, we discuss the potential of stalks to facilitate nutrient uptake and propose additional physiological roles for stalk elongation in Caulobacter cells.

Physiological role of stalk lengthening in Caulobacter crescentus.

The cyclic dinucleotide c-di-GMP is a signaling molecule with diverse functions in cellular physiology. Here, we report that c-di-GMP can assemble into a tetramer that mediates the effective dimerization of a transcription factor, BldD, which controls the progression of multicellular differentiation in sporulating actinomycete bacteria. BldD represses expression of sporulation genes during vegetative growth in a manner that depends on c-di-GMP-mediated dimerization. Structural and biochemical analyses show that tetrameric c-di-GMP links two subunits of BldD through their C-terminal domains, which are otherwise separated by ~10 Å and thus cannot effect dimerization directly. Binding of the c-di-GMP tetramer by BldD is selective and requires a bipartite RXD-X8-RXXD signature. The findings indicate a unique mechanism of protein dimerization and the ability of nucleotide signaling molecules to assume alternative oligomeric states to effect different functions.

Tetrameric c-di-GMP mediates effective transcription factor dimerization to control Streptomyces development.

The alphaproteobacterium Hyphomonas neptunium proliferates by a unique budding mechanism in which daughter cells emerge from the end of a stalk-like extension emanating from the mother cell body. Studies of this species so far have been hampered by the lack of a genetic system and of molecular tools allowing the regulated expression of target genes. Based on microarray analyses, this work identifies two H. neptunium promoters that are activated specifically by copper and zinc. Functional analyses show that they have low basal activity and a high dynamic range, meeting the requirements for use as a multipurpose expression system. To facilitate their application, the two promoters were incorporated into a set of integrative plasmids, featuring a choice of two different selection markers and various fluorescent protein genes. These constructs enable the straightforward generation and heavy metal-inducible synthesis of fluorescent protein fusions in H. neptunium, thereby opening the door to an in-depth analysis of polar growth and development in this species.

Molecular toolbox for genetic manipulation of the stalked budding bacterium Hyphomonas neptunium.

The complex life cycle of streptomycetes involves two distinct filamentous cell forms: the growing (or vegetative) hyphae and the reproductive (or aerial) hyphae, which differentiate into long chains of spores. Until recently, little was known about the signalling pathways that regulate the developmental transitions leading to sporulation. In this Review, we discuss important new insights into these pathways that have led to the emergence of a coherent regulatory network, focusing on the erection of aerial hyphae and the synchronous cell division event that produces dozens of unigenomic spores. In particular, we highlight the role of cyclic di-GMP (c-di-GMP) in controlling the initiation of development, and the role of the master regulator BldD in mediating c-di-GMP signalling.

Susan Schlimpert

Department of Molecular Microbiology

John Innes Centre, Norwich Research Park

Fluorescence Time-lapse Imaging of the Complete S. venezuelae Life Cycle Using a Microfluidic Device

Susan Schlimpert

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Department of Molecular Microbiology

John Innes Centre, Norwich Research Park

Fluorescence Time-lapse Imaging of the Complete S. venezuelae Life Cycle Using a Microfluidic Device

Department of Molecular Microbiology