the-c-elegans-intestine-as-model-for-intercellular-lumen

The C. elegans Intestine As a Model for Intercellular Lumen Morphogenesis and In Vivo Polarized Membrane Biogenesis at the Single-cell Level: Labeling by Antibody Staining, RNAi Loss-of-function Analysis and Imaging

Multicellular tubes, fundamental units of all internal organs, are composed of polarized epithelial or endothelial cells, with apical membranes lining the ...

the-c-elegans-excretory-canal-as-model-for-intracellular-lumen

The C. elegans Excretory Canal as a Model for Intracellular Lumen Morphogenesis and In Vivo Polarized Membrane Biogenesis in a Single Cell: labeling by GFP-fusions, RNAi Interaction Screen and Imaging

The four C. elegans excretory canals are narrow tubes extended through the length of the animal from a single cell, with almost equally far extended ...

Harvard Medical School

The extent to which excitable cells and behavior modulate animal development has not been examined in detail. Here, we demonstrate the existence of a novel pathway for promoting vulval fates in C. elegans that involves activation of the heterotrimeric Galphaq protein, EGL-30. EGL-30 acts with muscle-expressed EGL-19 L-type voltage-gated calcium channels to promote vulva development, and acts downstream or parallel to LET-60 (RAS). This pathway is not essential for vulval induction on standard Petri plates, but can be stimulated by expression of activated EGL-30 in neurons, or by an EGL-30-dependent change in behavior that occurs in a liquid environment. Our results indicate that excitable cells and animal behavior can provide modulatory inputs into the effects of growth factor signaling on cell fates, and suggest that communication between these cell populations is important for normal development to occur under certain environmental conditions.

Modulation of EGF receptor-mediated vulva development by the heterotrimeric G-protein Galphaq and excitable cells in C. elegans.

A recently identified molecule C-terminus of Hsc70 interacting protein (CHIP) has been reported to be an E3 ubiquitin ligase collaborating with molecular chaperones for the degradation of misfolded or unfolded proteins. The physiological roles of CHIP in animal and plant development remain largely unknown. Here, we show that the knockdown of CeCHIP by RNAi and knockout by a deletion mutation arrests the development of the animal at the larval stage. CeCHIP expresses ubiquitously in all tissues but there are tissue specific variations of expression level. CeCHIP produces dose dependent phenotypes in vivo. Over expression of CHIP causes embryonic lethality, while a comparatively lower level of over expression causes locomotion and egg laying defects, and the CHIP over expressed animals form dauers at a higher temperature.

Molecular and functional analysis of Caenorhabditis elegans CHIP, a homologue of Mammalian CHIP.

Polyglutamine (polyQ) expansion in many proteins, including huntingtin and ataxin-3, is pathogenic and responsible for neuronal dysfunction and degeneration. Although at least nine neurodegenerative diseases are caused by expanded polyQ, the pathogenesis of these diseases is still not well understood. In the present study, we used Caenorhabditis elegans to study the molecular mechanism of polyQ-mediated toxicity. We expressed full-length and truncated ataxin-3 with different lengths of polyQ in the nervous system of C. elegans. We show that expanded polyQ interrupts synaptic transmission, and induces swelling and aberrant branching of neuronal processes. Using an ubiquitinated fluorescence reporter construct, we also showed that polyQ aggregates impair the ubiquitin-proteasome system in C. elegans. These results may provide information for further understanding the pathogenesis of polyQ diseases.

Expanded polyglutamines impair synaptic transmission and ubiquitin-proteasome system in Caenorhabditis elegans.

Neuronal homeostasis requires a balance between anabolic and catabolic processes. Eukaryotic cells use two distinct systems for the degradation of unused proteins: the ubiquitin-proteasome system and the autophagic system. The autophagic system is also necessary for the degradation of bulk amounts of proteins and organelles. We have searched for new autophagy-related genes in the Caenorhabditis elegans genome and investigated their role in a polyglutamine (polyQ) disease model. Here, we have shown that inactivation of these genes intensified the toxicity of expanded polyQ in C. elegans neurons and muscles, and at the same time inactivation of CeTor reduced the polyQ toxicity.

Genetic impairment of autophagy intensifies expanded polyglutamine toxicity in Caenorhabditis elegans.

Metazoan internal organs are assembled from polarized tubular epithelia that must set aside an apical membrane domain as a lumenal surface. In a global Caenorhabditis elegans tubulogenesis screen, interference with several distinct fatty-acid-biosynthetic enzymes transformed a contiguous central intestinal lumen into multiple ectopic lumens. We show that multiple-lumen formation is caused by apicobasal polarity conversion, and demonstrate that in situ modulation of lipid biosynthesis is sufficient to reversibly switch apical domain identities on growing membranes of single post-mitotic cells, shifting lumen positions. Follow-on targeted lipid-biosynthesis pathway screens and functional genetic assays were designed to identify a putative single causative lipid species. They demonstrate that fatty-acid biosynthesis affects polarity through sphingolipid synthesis, and reveal ceramide glucosyltransferases (CGTs) as end-point biosynthetic enzymes in this pathway. Our findings identify glycosphingolipids, CGT products and obligate membrane lipids, as critical determinants of in vivo polarity and indicate that they sort new components to the expanding apical membrane.

Apicobasal domain identities of expanding tubular membranes depend on glycosphingolipid biosynthesis.

Clathrin coats vesicles in all eukaryotic cells and has a well-defined role in endocytosis, moving molecules away from the plasma membrane. Its function on routes towards the plasma membrane was only recently appreciated and is thought to be limited to basolateral transport. Here, an unbiased RNAi-based tubulogenesis screen identifies a role of clathrin (CHC-1) and its AP-1 adaptor in apical polarity during de novo lumenal membrane biogenesis in the C. elegans intestine. We show that CHC-1/AP-1-mediated polarized transport intersects with a sphingolipid-dependent apical sorting process. Depleting each presumed trafficking component mislocalizes the same set of apical membrane molecules basolaterally, including the polarity regulator PAR-6, and generates ectopic lateral lumens. GFP::CHC-1 and BODIPY-ceramide vesicles associate perinuclearly and assemble asymmetrically at polarized plasma membrane domains in a co-dependent and AP-1-dependent manner. Based on these findings, we propose a trafficking pathway for apical membrane polarity and lumen morphogenesis that implies: (1) a clathrin/AP-1 function on an apically directed transport route; and (2) the convergence of this route with a sphingolipid-dependent apical trafficking path.

Clathrin and AP-1 regulate apical polarity and lumen formation during C. elegans tubulogenesis.

&#x8BB8;&#x591A;&#x5355;&#x7EC6;&#x80DE;&#x7BA1;&#x5982;&#x6BDB;&#x7EC6;&#x8840;&#x7BA1;&#x5F62;&#x6210;&#x6D41;&#x660E;&#x819C;&#x5C42;&#xFF0C;&#x4E00;&#x4E2A;&#x4E0D;&#x5F88;&#x4E86;&#x89E3;&#x7684;&#x8FC7;&#x7A0B;&#x3002;&#x5728;&#x8FD9;&#x91CC;&#x6211;&#x4EEC;&#x5C55;&#x793A; ERM-1 &#x76AE;&#x8D28;&#x819C;&#x7EC4;&#x7EC7;&#x8005;&#x9700;&#x5728;&#x5355;&#x4E2A;&#x5355;&#x5143;&#x683C;&#x79C0;&#x4E3D;&#x9690;&#x6746;&#x7EBF;&#x866B;&#x6392;&#x6CC4;&#x8FD0;&#x6CB3;&#x5F62;&#x6001;&#x53D1;&#x751F;&#x671F;&#x95F4;&#x5C55;&#x5F00;&#x7684;&#x80DE;&#x5185;&#x7684;&#x5FC3;&#x5C16;/&#x80A0;&#x58C1;&#x819C;&#x548C;&#x5176;&#x808C;&#x52A8;&#x86CB;&#x767D;&#x5185;&#x6D82;&#x5C42;&#x3002;&#x6211;&#x4EEC;&#x7684;&#x7279;&#x70B9;&#x6C34;&#x901A;&#x9053;&#x86CB;&#x767D;-8&#xFF0C;&#x786E;&#x5B9A;&#x7684; ERM 1 &#x8868;&#x8FBE; (ERM-1[++]) &#x6291;&#x5236;&#x5668;&#x5C4F;&#x5E55;&#x4E0A;&#xFF0C;&#x4F5C;&#x4E3A;&#x6CEA;&#x9053;&#x7684;&#x6C34;&#x901A;&#x9053;&#x86CB;&#x767D;&#x8FDB;&#x884C;&#x4EA4;&#x4E92;&#x4EE5; ERM-1 &#x6D41;&#x660E;&#x6269;&#x5C55;&#x4E2D;&#x6C5E;&#x654F;&#x611F;&#x7684;&#x65B9;&#x5F0F;&#xFF0C;&#x6C34;&#x901A;&#x9053;&#x6D3B;&#x52A8;&#x6709;&#x7275;&#x8FDE;&#x3002;&#x6C34;&#x901A;&#x9053;&#x86CB;&#x767D;-8 &#x662F;&#x77AC;&#x6001;&#x62DB;&#x52DF;&#x7531; ERM-1&#xFF0C;&#x5728;&#x56F4;&#x80A0;&#x58C1;&#x8896;&#x53E3;&#x6CBF;&#x6269;&#x5927;&#x8FD0;&#x6CB3; interspaced co-localizing &#x6D41;&#x660E;&#x3002;ERM-1 + [+]-&#x4ECB;&#x5BFC;&#x7684;&#x589E;&#x52A0;&#x6D41;&#x660E;&#x5173;&#x8054;&#x5C0F;&#x7BA1;&#x65AD;&#x88C2;&#x7684;&#x6570;&#x76EE;&#x626D;&#x8F6C;&#x7684;&#x6C34;&#x901A;&#x9053;&#x86CB;&#x767D; 8 &#x67AF;&#x7AED;&#x3002;&#x6211;&#x4EEC;&#x5EFA;&#x8BAE;&#x7531; translumenal &#x901A;&#x91CF;&#xFF0C;ERM-1/&#x6C34;&#x901A;&#x9053;&#x86CB;&#x767D;-8 &#x4E92;&#x52A8;&#x63A8;&#x8FDB;&#x6D41;&#x660E;&#x6269;&#x5C55;&#x540D;&#x6697;&#x793A;&#x7684;&#x6C34;&#x901A;&#x9053;&#x53D7;&#x89C4;&#x7BA1;&#x7684;&#x6D41;&#x4F53;&#x538B;&#x529B;&#x76F4;&#x63A5;&#x5F62;&#x6001;&#x53D1;&#x751F;&#x6548;&#x5E94;&#x3002;

&#x80DE;&#x8154;&#x6269;&#x5C55;&#x9700;&#x8981; ERM-1-&#x4F9D;&#x8D56;&#x9876;&#x7AEF;&#x819C;&#x6269;&#x5F20;&#x548C;&#x6C34;&#x901A;&#x9053;&#x86CB;&#x767D;&#x4ECB;&#x5BFC; 8 &#x901A;&#x91CF;&#x3002;

Many unicellular tubes such as capillaries form lumens intracellularly, a process that is not well understood. Here we show that the cortical membrane organizer ERM-1 is required to expand the intracellular apical/lumenal membrane and its actin undercoat during single-cell Caenorhabditis elegans excretory canal morphogenesis. We characterize AQP-8, identified in an ERM-1-overexpression (ERM-1[++]) suppressor screen, as a canalicular aquaporin that interacts with ERM-1 in lumen extension in a mercury-sensitive manner, implicating water-channel activity. AQP-8 is transiently recruited to the lumen by ERM-1, co-localizing in peri-lumenal cuffs interspaced along expanding canals. An ERM-1[++]-mediated increase in the number of lumen-associated canaliculi is reversed by AQP-8 depletion. We propose that the ERM-1/AQP-8 interaction propels lumen extension by translumenal flux, suggesting a direct morphogenetic effect of water-channel-regulated fluid pressure.

Intracellular lumen extension requires ERM-1-dependent apical membrane expansion and AQP-8-mediated flux.

Nombreux tubes unicellulaires comme les capillaires forment lumens intracellulaire, un processus qui n&#39;est pas bien compris. Ici, nous montrons que l&#39;organisateur de la membrane corticale ERM-1 est n&eacute;cessaire pour &eacute;largir la membrane apicale/lumi&egrave;re intracellulaire et son sous-poil actine au cours de la seule cellule Caenorhabditis elegans canal excr&eacute;teur morphogen&egrave;se. Nous caract&eacute;risons AQP-8, identifi&eacute; dans une MCE-1-surexpression (&eacute;cran de suppresseur ERM-1[++]), comme une aquaporine canaliculaire qui interagit avec MCE-1 en prolongation de lumen de mani&egrave;re sensible au mercure, impliquant l&#39;activit&eacute; de l&#39;eau-canal. AQP-8 est transitoirement recrut&eacute;s &agrave; la lumi&egrave;re de MCE-1, co-localizing en peri-lumi&egrave;re poignets espac&eacute;es le long de l&#39;expansion des canaux. Un MTC-1 [++]-m&eacute;di&eacute;e par augmentation du nombre des canalicules associ&eacute;s &agrave; lumen est renvers&eacute;e par l&#39;appauvrissement AQP-8. Nous proposons que l&#39;interaction de l&#39;ERM-1/AQP-8 propulse extension lumen par flux translumenal, sugg&eacute;rant un effet morphog&eacute;n&eacute;tique direct de pression du fluide eau-canal-r&eacute;gl&eacute;es.

Lumen intracellulaire extension n&eacute;cessite expansion membrane apicale MCE-1-d&eacute;pendante et le flux induite par l&#39;AQP-8.

Molti tubi unicellulari come capillari formano lumen intracellularly, un processo che non &egrave; ben compreso. Qui ci mostra che l&#39;organizzatore della membrana corticale ERM-1 &egrave; necessario espandere la membrana apicale/lumenal intracellulare e il suo sottopelo actina durante la cella singola Caenorhabditis elegans canale escretore morfogenesi. Ci caratterizzano AQP-8, identificato in una sovraespressione ERM-1 (ERM-1[++]) schermo soppressore, come un aquaporina canalicolare che interagisce con ERM-1 nell&#39;estensione del lume in maniera sensibile al mercurio, implicando attivit&agrave; di acqua-canale. AQP-8 &egrave; transitoriamente reclutati per il lume di ERM-1, co-localizing in peri-lumenal polsini intervallate lungo canali in espansione. Un ERM-1 [+ +]-mediata aumento del numero di associati al lume canaliculi &egrave; invertito da deplezione di AQP-8. Proponiamo che l&#39;interazione di ERM-1/AQP-8 aziona estensione lumen di flusso translumenal, suggerendo un effetto morfogenetico diretto della pressione del fluido acqua-canale-regolato.

Lume intracellulare estensione richiede espansione membrana apicale ERM-1-dipendente e flux AQP-8-mediata.

&#x6BDB;&#x7D30;&#x8840;&#x7BA1;&#x306A;&#x3069;&#x591A;&#x304F;&#x306E;&#x5358;&#x7D30;&#x80DE;&#x7BA1;&#x5185;&#x8154;&#x7D30;&#x80DE;&#x5185;&#x3001;&#x30D5;&#x30A9;&#x30FC;&#x30E0;&#x3088;&#x304F;&#x7406;&#x89E3;&#x3055;&#x308C;&#x3066;&#x3044;&#x306A;&#x3044;&#x30D7;&#x30ED;&#x30BB;&#x30B9;&#x3002;&#x3053;&#x3053;&#x3067;&#x76AE;&#x8CEA;&#x819C;&#x30AA;&#x30FC;&#x30AC;&#x30CA;&#x30A4;&#x30B6;&#x30FC; ERM 1 &#x7D30;&#x80DE;&#x9802;/&#x306A;&#x3063;&#x305F;&#x5185;&#x8154;&#x5074;&#x819C;&#x3068;&#x305D;&#x306E;&#x30A2;&#x30AF;&#x30C1;&#x30F3;&#x4E0B;&#x5730;&#x5358;&#x4E00;&#x30BB;&#x30EB;&#x7DDA;&#x866B;&#x7DDA;&#x866B;&#x6392;&#x6CC4;&#x904B;&#x6CB3;&#x306E;&#x5F62;&#x614B;&#x5F62;&#x6210;&#x4E2D;&#x306B;&#x5C55;&#x958B;&#x3059;&#x308B;&#x5FC5;&#x8981;&#x304C;&#x3042;&#x308B;&#x3053;&#x3068;&#x3092;&#x793A;&#x3057;&#x307E;&#x3059;&#x3002;&#x30A2;&#x30AF;&#x30A2;&#x30DD;&#x30EA;&#x30F3;-8&#x3001;ERM-1 &#x767A;&#x73FE; (ERM-1[++]) &#x6291;&#x5236;&#x753B;&#x9762;&#x3001;ERM-1 &#x3068;&#x306E;&#x5185;&#x8154;&#x62E1;&#x5F35; implicating &#x6C34;&#x30C1;&#x30E3;&#x30CD;&#x30EB;&#x306E;&#x5229;&#x7528;&#x72B6;&#x6CC1;&#x3001;&#x6C34;&#x9280;&#x306B;&#x654F;&#x611F;&#x306A;&#x65B9;&#x6CD5;&#x3067;&#x5BFE;&#x8A71;&#x6BDB;&#x7D30;&#x30A2;&#x30AF;&#x30A2;&#x30DD;&#x30EA;&#x30F3;&#x3068;&#x3057;&#x3066;&#x3002; &#x8B58;&#x5225;&#x3092;&#x7279;&#x5FB4;&#x4ED8;&#x3051;&#x308B;&#x30A2;&#x30AF;&#x30A2;&#x30DD;&#x30EA;&#x30F3; 8 &#x306F;&#x4E00;&#x904E;&#x6027;&#x306B;&#x5185;&#x8154; ERM - 1 co-localizing &#x904B;&#x6CB3;&#x3092;&#x62E1;&#x5927;&#x306B;&#x6CBF;&#x3063;&#x3066; interspaced &#x30DA;&#x30EA;&#x306A;&#x3063;&#x305F;&#x5185;&#x8154;&#x5074;&#x306E;&#x8896;&#x53E3;&#x3067;&#x52DF;&#x96C6;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;ERM 1 &#x306E; [+ +]-&#x7D30;&#x7BA1;&#x306E;&#x5185;&#x8154;&#x306B;&#x95A2;&#x9023;&#x4ED8;&#x3051;&#x3089;&#x308C;&#x305F;&#x6570;&#x306E;&#x5A92;&#x4ECB;&#x5897;&#x52A0;&#x304C;&#x9006;&#x30A2;&#x30AF;&#x30A2;&#x30DD;&#x30EA;&#x30F3; 8 &#x6E1B;&#x5C11;&#x306B;&#x3088;&#x3063;&#x3066;&#x3002;&#x6211;&#x3005; &#x306F;&#x6C34;&#x30C1;&#x30E3;&#x30CD;&#x30EB;&#x8ABF;&#x6574;&#x5727;&#x529B;&#x306E;&#x76F4;&#x63A5;&#x5F62;&#x6210;&#x52B9;&#x679C;&#x3092;&#x793A;&#x5506;&#x3001;ERM 1/&#x30A2;&#x30AF;&#x30A2;&#x30DD;&#x30EA;&#x30F3; 8 &#x76F8;&#x4E92;&#x4F5C;&#x7528;&#x304C;&#x8D8A;&#x7D4C;&#x7BA1;&#x8154;&#x30D5;&#x30E9;&#x30C3;&#x30AF;&#x30B9;&#x306B;&#x3088;&#x308B;&#x5185;&#x8154;&#x62E1;&#x5F35;&#x3092;&#x63A8;&#x9032;&#x3059;&#x308B;&#x3053;&#x3068;&#x3092;&#x63D0;&#x6848;&#x3057;&#x307E;&#x3059;&#x3002;

&#x7D30;&#x80DE;&#x5185;&#x306E;&#x5185;&#x8154;&#x62E1;&#x5F35;&#x6A5F;&#x80FD;&#x3067;&#x306F;&#x3001;ERM-1 &#x4F9D;&#x5B58;&#x30A2;&#x30D4;&#x30AB;&#x30EB;&#x819C;&#x306E;&#x62E1;&#x5F35;&#x30A2;&#x30AF;&#x30A2;&#x30DD;&#x30EA;&#x30F3; 8 &#x3092;&#x4ECB;&#x3057;&#x305F;&#x30D5;&#x30E9;&#x30C3;&#x30AF;&#x30B9;&#x3092;&#x5FC5;&#x8981;&#x304C;&#x3042;&#x308A;&#x307E;&#x3059;&#x3002;

&#xBAA8;&#xC138; &#xD608;&#xAD00; &#xB4F1; &#xB9CE;&#xC740; &#xB2E8; &#xC138;&#xD3EC; &#xAD00; &#xCE68;, &#xB8E8;&#xBA58;&#xC744; &#xD615;&#xC131; &#xD504;&#xB85C;&#xC138;&#xC2A4;&#xB97C; &#xC798; &#xC774;&#xD574; &#xB418;&#xC9C0; &#xC54A;&#xC2B5;&#xB2C8;&#xB2E4;. &#xC5EC;&#xAE30;&#xC5D0; &#xC6B0;&#xB9AC;&#xAC00; &#xD45C;&#xC2DC; &#xC678;&#xD53C; &#xB9C9; organizer &#xC74C; 1 &#xB2E8;&#xC77C; &#xC140; &#xAF2C;&#xB9C8; &#xC120; &#xCDA9; &#xBC30;&#xC124; &#xC6B4;&#xD558; morphogenesis &#xB3D9;&#xC548; &#xC138;&#xD3EC;&#xB0B4; &#xAF2D;&#xB300;&#xAE30;/lumenal &#xB9C9;&#xACFC; &#xADF8;&#xAC83;&#xC758; &#xAC78; &#xC18D; &#xD138;&#xC744; &#xD655;&#xC7A5; &#xD558;&#xB294; &#xB370; &#xD544;&#xC694;&#xD55C;&#xC785;&#xB2C8;&#xB2E4;. &#xC6B0;&#xB9AC;&#xB294; &#xC74C;-1-overexpression (ERM-1[++]) &#xC5B5;&#xC555; &#xD654;&#xBA74;, &#xC74C; 1 &#xB8E8;&#xBA58; &#xD655;&#xC7A5;&#xC5D0;&#xC11C; &#xBB3C; &#xCC44;&#xB110; &#xD65C;&#xB3D9;&#xC744; &#xB0B4;&#xD3EC; &#xC218;&#xC740; &#xBBFC;&#xAC10;&#xD55C; &#xBC29;&#xC2DD;&#xC73C;&#xB85C; &#xC0C1;&#xD638;&#xC791;&#xC6A9; canalicular aquaporin&#xB85C; &#xC2DD;&#xBCC4; AQP 8 &#xD2B9;&#xC9D5;. &#xC815;&#xB3C4; &#xC74C;-1, peri lumenal &#xC218; &#xAC11; &#xC6B4;&#xD558; &#xD655;&#xC7A5; &#xD568;&#xAED8; interspaced&#xC5D0; co-localizing&#xC5D0; &#xC758;&#xD574; AQP 8 &#xB8E8;&#xBA58;&#xC744; &#xBAA8;&#xC9D1;. &#xC74C; 1 + [+]-&#xB8E8;&#xBA58; &#xAD00;&#xB828; canaliculi &#xC218; &#xC788;&#xB294; &#xC911;&#xC7AC; &#xC99D;&#xAC00; AQP 8 &#xACE0;&#xAC08;&#xC5D0; &#xC758;&#xD574; &#xBC18;&#xB300;&#xC785;&#xB2C8;&#xB2E4;. &#xC6B0;&#xB9AC; &#xC74C; 1/AQP 8 &#xC0C1;&#xD638; &#xC791;&#xC6A9; translumenal &#xD50C;&#xB7ED;&#xC2A4;, &#xB8E8;&#xBA58; &#xD655;&#xC7A5; &#xCD94;&#xC9C4; &#xBB3C; &#xCC44;&#xB110; &#xADDC;&#xC81C; &#xC720;&#xCCB4; &#xC555;&#xB825;&#xC758; &#xC9C1;&#xC811;&#xC801;&#xC778; &#xC804;&#xCCB4;&#xC801; &#xD6A8;&#xACFC; &#xC81C;&#xC548; &#xC81C;&#xC548; &#xD569;&#xB2C8;&#xB2E4;.

&#xC138;&#xD3EC;&#xB0B4; &#xB8E8;&#xBA58; &#xD655;&#xC7A5; &#xC74C; 1 &#xC885;&#xC18D; &#xAF2D;&#xB300;&#xAE30; &#xB9C9; &#xD655;&#xC7A5; AQP 8 &#xC911;&#xC7AC; &#xD558;&#xB294; &#xD50C;&#xB7ED;&#xC2A4;&#xB97C; &#xD544;&#xC694;&#xD569;&#xB2C8;&#xB2E4;.

Muchos tubos unicelulares como capilares forman l&uacute;menes intracelular, un proceso que no se comprende bien. Aqu&iacute; mostramos que el organizador de la membrana cortical ERM-1 es necesaria para ampliar la membrana apical/lumenal intracelular y su capa de actina durante la morfog&eacute;nesis de canal excretor unicelular Caenorhabditis elegans. Caracterizamos AQP-8, identificado en una sobreexpresi&oacute;n ERM-1 (pantalla del supresor ERM-1[++]), como un aquaporin con canal&iacute;culos que interact&uacute;a con ERM-1 extensi&oacute;n de luz de manera sensible al mercurio, implicando la actividad del canal de agua. AQP-8 es reclutado transitoriamente a la luz por ERM-1, Junta en pu&ntilde;os peri lumenal otro a lo largo de la expansi&oacute;n de canales. Un ERM-1 [++]-mediada por aumento en el n&uacute;mero de asociados de lumen canal&iacute;culos se invierte por el agotamiento de AQP-8. Proponemos que la interacci&oacute;n de ERM-1/AQP-8 impulsa la extensi&oacute;n de lumen por flujo translumenal, sugiriendo un efecto morfogen&eacute;tico directo de presi&oacute;n regulada por el canal de agua.

Lumen intracelular extensi&oacute;n requiere expansi&oacute;n de membrana apical de ERM-1-dependiente y AQP-8-mediada por flujo.

Biological tubes consist of polarized epithelial cells with apical membranes building the central lumen and basolateral membranes contacting adjacent cells or the extracellular matrix. Cellular polarity requires distinct inputs from outside the cell, e.g., the matrix, inside the cell, e.g., vesicular trafficking and the plasma membrane and its junctions.(1) Many highly conserved polarity cues have been identified, but their integration during the complex process of polarized tissue and organ morphogenesis is not well understood. It is assumed that plasma-membrane-associated polarity determinants, such as the partitioning-defective (PAR) complex, define plasma membrane domain identities, whereas vesicular trafficking delivers membrane components to these domains, but lacks the ability to define them. In vitro studies on lumenal membrane biogenesis in mammalian cell lines now indicate that trafficking could contribute to defining membrane domains by targeting the polarity determinants, e.g., the PARs, themselves.(2) This possibility suggests a mechanism for PARs' asymmetric distribution on membranes and places vesicle-associated polarity cues upstream of membrane-associated polarity determinants. In such an upstream position, trafficking might even direct multiple membrane components, not only polarity determinants, an original concept of polarized plasma membrane biogenesis(3) (,) (4)that was largely abandoned due to the failure to identify a molecularly defined intrinsic vesicular sorting mechanism. Our two recent studies on C. elegans intestinal tubulogenesis reveal that glycosphingolipids (GSLs) and the well-recognized vesicle components clathrin and its AP-1 adaptor are required for targeting multiple apical molecules, including polarity regulators, to the expanding apical/lumenal membrane.(5) (,) (6) These findings support GSLs' long-proposed role in in vivo polarized epithelial membrane biogenesis and development and identify a novel function in apical polarity for classical post-Golgi vesicle components. They are also compatible with a vesicle-intrinsic sorting mechanism during membrane biogenesis and suggest a model for how vesicles could acquire apical directionality during the assembly of the functionally critical polarized lumenal surfaces of epithelial tubes.

Vesicular sorting controls the polarity of expanding membranes in the C. elegans intestine.

The field of metabolomics continues to catalog new compounds, but their functional analysis remains technically challenging, and roles beyond metabolism are largely unknown. Unbiased genetic/RNAi screens are powerful tools to identify the in vivo functions of protein-encoding genes, but not of nonproteinaceous compounds such as lipids. They can, however, identify the biosynthetic enzymes of these compounds-findings that are usually dismissed, as these typically synthesize multiple products. Here, we provide a method using follow-on biosynthetic pathway screens to identify the endpoint biosynthetic enzyme and thus the compound through which they act. The approach is based on the principle that all subsequently identified downstream biosynthetic enzymes contribute to the synthesis of at least this one end product. We describe how to systematically target lipid biosynthetic pathways; optimize targeting conditions; take advantage of pathway branchpoints; and validate results by genetic assays and biochemical analyses. This approach extends the power of unbiased genetic/RNAi screens to identify in vivo functions of non-nucleic acid-based metabolites beyond their metabolic roles. It will typically require several months to identify a metabolic end product by biosynthetic pathway screens, but this time will vary widely depending, among other factors, on the end product's location in the pathway, which determines the number of screens required for its identification.

RNAi-based biosynthetic pathway screens to identify in vivo functions of non-nucleic acid-based metabolites such as lipids.

Corrigendum: RNAi-based biosynthetic pathway screens to identify in vivo functions of non-nucleic acid-based metabolites such as lipids.

Determination of luminal diameter is critical to the function of small single-celled tubes. A series of EXC proteins, including EXC-1, prevent swelling of the tubular excretory canals in Caenorhabditis elegans In this study, cloning of exc-1 reveals it to encode a homolog of mammalian IRG proteins, which play roles in immune response and autophagy and are associated with Crohn's disease. Mutants in exc-1 accumulate early endosomes, lack recycling endosomes, and exhibit abnormal apical cytoskeletal structure in regions of enlarged tubules. EXC-1 interacts genetically with two other EXC proteins that also affect endosomal trafficking. In yeast two-hybrid assays, wild-type and putative constitutively active EXC-1 binds to the LIM-domain protein EXC-9, whose homolog, cysteine-rich intestinal protein, is enriched in mammalian intestine. These results suggest a model for IRG function in forming and maintaining apical tubule structure via regulation of endosomal recycling.

ArticleTotal : 2	Year
Publication title Cited by 12	2017
Publication title Cited by 6	2017

Article	Year
Modulation of EGF receptor-mediated vulva development by the heterotrimeric G-protein Galphaq and excitable cells in C. elegans. Development (Cambridge, England)\| PubMed ID: 12925583	2003
Molecular and functional analysis of Caenorhabditis elegans CHIP, a homologue of Mammalian CHIP. FEBS letters\| PubMed ID: 15135044	2004
Expanded polyglutamines impair synaptic transmission and ubiquitin-proteasome system in Caenorhabditis elegans. Journal of neurochemistry\| PubMed ID: 16805848	2006
Genetic impairment of autophagy intensifies expanded polyglutamine toxicity in Caenorhabditis elegans. Biochemical and biophysical research communications\| PubMed ID: 18261464	2008
Apicobasal domain identities of expanding tubular membranes depend on glycosphingolipid biosynthesis. Nature cell biology\| PubMed ID: 21926990	2011
Clathrin and AP-1 regulate apical polarity and lumen formation during C. elegans tubulogenesis. Development (Cambridge, England)\| PubMed ID: 22535410	2012
Intracellular lumen extension requires ERM-1-dependent apical membrane expansion and AQP-8-mediated flux. Nature cell biology\| PubMed ID: 23334498	2013
Vesicular sorting controls the polarity of expanding membranes in the C. elegans intestine. Worm\| PubMed ID: 24058862	2013
RNAi-based biosynthetic pathway screens to identify in vivo functions of non-nucleic acid-based metabolites such as lipids. Nature protocols\| PubMed ID: 25837419	2015
Corrigendum: RNAi-based biosynthetic pathway screens to identify in vivo functions of non-nucleic acid-based metabolites such as lipids. Nature protocols\| PubMed ID: 26020617	2015
Facilitation of Endosomal Recycling by an IRG Protein Homolog Maintains Apical Tubule Structure in Caenorhabditis elegans. Genetics\| PubMed ID: 27334269	2016

Liakot A Khan

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