high-resolution-whole-mount-situ-hybridization-within-zebrafish

High Resolution Whole Mount <em>In Situ</em> Hybridization within Zebrafish Embryos to Study Gene Expression and Function

This article focuses on whole-mount in situ hybridization (WISH) of zebrafish embryos. The WISH technology facilitates the assessment of gene expression ...

Royal Victoria Hospital

McGill University Health Centre Research Institute

Carp granulins are members of an emerging class of proteins with a sequence motif encoding a parallel stack of two to four beta-hairpins. The carp granulin-1 protein forms a stack of four beta-hairpins, whereas its amino-terminal fragment appears to adopt a very stable stack of two beta-hairpins in solution. Here we determined a refined three-dimensional structure of this peptide fragment to examine potential conformational changes compared with the full-length protein. The structures were calculated with both a traditional method and a fast semiautomated method using ambiguous NMR distance restraints. The resulting sets of structures are very similar and show that a well-defined stack of two beta-hairpins is retained in the peptide. Conformational rearrangements compensating the loss of the carboxy-terminal subdomain of the native protein are restricted to the carboxy-terminal end of the peptide, the turn connecting the two beta-hairpins, and the Tyr(21) and Tyr(25) aromatic side chains. Further removal of the Val(1) and Ile(2) residues, which are part of the first beta-hairpin and components of two major hydrophobic clusters in the two beta-hairpin structure, results in the loss of the first beta-hairpin. The second beta-hairpin, which is closely associated with the first, retains a similar but somewhat less stable conformation. The invariable presence of the second beta-hairpin and the dependence of its stability on the first beta-hairpin suggest that the stack of two beta-hairpins may be an evolutionary conserved and autonomous folding unit. In addition, the high conformational stability makes the stack of two beta-hairpins an attractive scaffold for the development of peptide-based drug candidates.

Solution structures of a 30-residue amino-terminal domain of the carp granulin-1 protein and its amino-terminally truncated 3-30 subfragment: implications for the conformational stability of the stack of two beta-hairpins.

Progranulin is an epithelial tissue growth factor (also known as proepithelin, acrogranin and PC-cell-derived growth factor) that has been implicated in development, wound healing and in the progression of many cancers. The single mammalian progranulin gene encodes a glycoprotein precursor consisting of seven and one half tandemly repeated non-identical copies of the cystine-rich granulin motif. A genome-wide duplication event hypothesized to have occurred at the base of the teleost radiation predicts that mammalian progranulin may be represented by two co-orthologues in zebrafish.

The zebrafish progranulin gene family and antisense transcripts.

&#x795E;&#x7ECF;&#x8C03;&#x8282;&#x7EC6;&#x80DE;&#x51CB;&#x4EA1; convertase-1/&#x7EC6;&#x83CC;&#x6027; convertase &#x67AF;&#x8349;&#x79D1;&#x4FE1;&#x7C7B;&#x4F3C;-9 &#xFF08;&#x540C;-1/PCSK9) &#x662F;&#x7EC6;&#x83CC;&#x6027;&#x7684; convertase &#x6700;&#x8FD1;&#x63CF;&#x8FF0;&#x4E86;&#x901A;&#x8FC7;&#x589E;&#x5F3A;&#x4F4E;&#x5BC6;&#x5EA6;&#x8102;&#x86CB;&#x767D;&#x53D7;&#x4F53; (LDLR) &#x964D;&#x89E3;&#x80C6;&#x56FA;&#x9187;&#x7A33;&#x6001;&#x548C;&#x53EF;&#x80FD;&#x5728;&#x795E;&#x7ECF;&#x53D1;&#x80B2;&#x4E2D;&#x53D1;&#x6325;&#x4E3B;&#x8981;&#x4F5C;&#x7528;&#x3002;&#x672C;&#x6587;&#x6240;&#x8FF0;&#xFF0C;&#x6211;&#x4EEC;&#x8C03;&#x67E5;&#x4E86;&#x8FD9;&#x86CB;&#x767D;&#x9176;&#x4F5C;&#x4E3A;&#x6A21;&#x578B;&#x4F7F;&#x7528;&#x9F20;&#x6807; P19 &#x80DA;&#x80CE;&#x591A;&#x80FD;&#x5E72;&#x7EC6;&#x80DE;&#x548C;&#x6591;&#x9A6C;&#x9C7C;&#x4E2D;&#x67A2;&#x795E;&#x7ECF;&#x7CFB;&#x7EDF;&#x53D1;&#x5C55;&#x4E2D;&#x7684;&#x6F5C;&#x5728;&#x53C2;&#x4E0E;&#x3002;&#x65F6;&#x95F4;&#x8BFE;&#x7A0B;&#x5B9A;&#x91CF;&#x9006;&#x8F6C;&#x5F55;-&#x805A;&#x5408;&#x9176;&#x94FE;&#x53CD;&#x5E94;&#x5206;&#x6790;&#x88AB;&#x6267;&#x884C;&#x4E4B;&#x540E;&#x89C6;&#x9EC4;&#x9178; (RA)-&#x8BF1;&#x5BFC; P19 &#x7EC6;&#x80DE;&#x795E;&#x7ECF;&#x5916;&#x80DA;&#x5C42;&#x5206;&#x5316;&#x3002;&#x56E0;&#x6B64;&#xFF0C;&#x540C;-1/PCSK9 &#x7684; mRNA &#x6C34;&#x5E73;&#x5728;&#x5206;&#x5316;&#x7684;&#x7B2C; 2 &#x5929;&#x8FBE;&#x5230;&#x6700;&#x9AD8;&#x5CF0;&#xFF0C;&#x6B64;&#x540E;&#x6389;&#x4E0B;&#x6765;&#x4E86;&#x3002;&#x76F8;&#x6BD4;&#x4E4B;&#x4E0B;&#xFF0C;&#x7EC6;&#x83CC;&#x6027; convertases &#x67AF;&#x8349;&#x79D1;&#x4FE1;&#x540C;&#x5DE5;&#x9176; 1/&#x7AD9;&#x70B9; 1 &#x86CB;&#x767D;&#x9176;&#x548C;&#x5F17;&#x6797;&#x86CB;&#x767D;&#x9176;&#x7684;&#x8868;&#x8FBE;&#x662F;&#x53D7; RA&#xFF0C;&#x800C; PC5/6 &#x548C; PC2 &#x5206;&#x522B;&#x589E;&#x52A0;&#x5185;&#x548C; (&#x6216;) &#x540E;&#x7B2C; 4 &#x5929;&#x7684;&#x5206;&#x5316;&#x671F;&#x3002;&#x8FD9;&#x79CD;&#x6A21;&#x5F0F;&#x5E76;&#x4E0D;&#x53D7; cholesterogenic &#x8F6C;&#x5F55;&#x56E0;&#x5B50;&#x753E;&#x9187;&#x89C4;&#x7BA1;&#x5143;&#x7D20;&#x7ED1;&#x5B9A;&#x86CB;&#x767D;-2&#xFF0C;&#x5176;&#x4E2D;&#x901A;&#x5E38;&#x5411;&#x4E0A;&#x8C03;&#x8282;&#x809D;&#x540C;-1/PCSK9 mRNA &#x6C34;&#x5E73;&#x3002;&#x6B64;&#x5916;&#xFF0C;&#x5728; P19 &#x7EC6;&#x80DE;&#xFF0C;RA &#x7684;&#x5F85;&#x9047;&#x5E76;&#x4E0D;&#x5F71;&#x54CD;&#x5185;&#x6E90;&#x6027;&#x7684; LDLR &#x86CB;&#x767D;&#x8D28;&#x6C34;&#x5E73;&#x3002;&#x8FD9;&#x540C;&#x610F;&#x540C;-1/PCSK9 &#x7684;&#x4E2D;&#x67A2;&#x795E;&#x7ECF;&#x7CFB;&#x7EDF;&#xFF0C;&#x5305;&#x62EC;&#x7684;&#x5C0F;&#x8111;&#xFF0C;&#x6CA1;&#x6709;&#x660E;&#x663E;&#x8868;&#x793A; LDLR &#x556E;&#x9F7F;&#x7C7B;&#x52A8;&#x7269;&#x4E2D;&#x7684;&#x72EC;&#x7279;&#x8868;&#x8FBE;&#x6A21;&#x5F0F;&#x3002;&#x6574;&#x4E2A;&#x88C5;&#x8F7D;&#x539F;&#x4F4D;&#x6742;&#x4EA4;&#x63ED;&#x793A;&#x7684;&#x6591;&#x9A6C;&#x9C7C;&#x7684;&#x8868;&#x8FBE;&#x6A21;&#x5F0F;&#x540C;-1/PCSK9 &#x662F;&#x7C7B;&#x4F3C;&#x4E8E;&#x9F20;&#x6807;&#x5728;&#x4E2D;&#x67A2;&#x795E;&#x7ECF;&#x7CFB;&#x7EDF;&#x548C;&#x5916;&#x56F4;&#x3002;&#x6591;&#x9A6C;&#x9C7C;&#x540C;-1/PCSK9 mRNA &#x7684;&#x7279;&#x5B9A; &#x2477; &#x9020;&#x6210;&#x5C0F;&#x8111;&#x795E;&#x7ECF;&#x7EC6;&#x80DE;&#x7684;&#x4F5C;&#x7528;&#x4E00;&#x822C;&#x89E3;&#x4F53;&#x548C;&#x540E;&#x8111;&#x4E2D;&#x8111;&#x7684;&#x754C;&#x9650;&#xFF0C;&#x5BFC;&#x81F4;&#x80DA;&#x80CE;&#x6B7B;&#x4EA1;&#x7EA6; 96 h &#x5728;&#x53D7;&#x7CBE;&#x540E;&#x7684;&#x635F;&#x5931;&#x3002;&#x8FD9;&#x4E9B;&#x6570;&#x636E;&#x540C;-1/PCSK9 &#x652F;&#x6301;&#x65B0;&#x578B;&#x7684;&#x4F5C;&#x7528;&#x5728;&#x4E2D;&#x67A2;&#x795E;&#x7ECF;&#x7CFB;&#x7EDF;&#x53D1;&#x80B2;&#xFF0C;&#x6709;&#x522B;&#x4E8E; cholesterogenic &#x5668;&#x5B98;&#x5982;&#x809D;&#x810F;&#x4E2D;&#x3002;

&#x7EC6;&#x83CC;&#x6027; convertase &#x540C;-1/PCSK9 &#x4E2D;&#x67A2;&#x795E;&#x7ECF;&#x7CFB;&#x7EDF;&#x53D1;&#x5C55;&#x4E2D;&#x7684;&#x610F;&#x4E49;&#x3002;

Neural apoptosis-regulated convertase-1/proprotein convertase subtilisin-kexin like-9 (NARC-1/PCSK9) is a proprotein convertase recently described to play a major role in cholesterol homeostasis through enhanced degradation of the low-density lipoprotein receptor (LDLR) and possibly in neural development. Herein, we investigated the potential involvement of this proteinase in the development of the CNS using mouse embryonal pluripotent P19 cells and the zebrafish as models. Time course quantitative RT-PCR analyses were performed following retinoic acid (RA)-induced neuroectodermal differentiation of P19 cells. Accordingly, the mRNA levels of NARC-1/PCSK9 peaked at day 2 of differentiation and fell off thereafter. In contrast, the expression of the proprotein convertases subtilisin kexin isozyme 1/site 1 protease and Furin was unaffected by RA, whereas that of PC5/6 and PC2 increased within and/or after the first 4 days of the differentiation period respectively. This pattern was not affected by the cholesterogenic transcription factor sterol regulatory element-binding protein-2, which normally up-regulates NARC-1/PCSK9 mRNA levels in liver. Furthermore, in P19 cells, RA treatment did not affect the protein level of the endogenous LDLR. This agrees with the unique expression pattern of NARC-1/PCSK9 in the rodent CNS, including the cerebellum, where the LDLR is not significantly expressed. Whole-mount in situ hybridization revealed that the pattern of expression of zebrafish NARC-1/PCSK9 is similar to that of mouse both in the CNS and periphery. Specific knockdown of zebrafish NARC-1/PCSK9 mRNA resulted in a general disorganization of cerebellar neurons and loss of hindbrain-midbrain boundaries, leading to embryonic death at approximately 96 h after fertilization. These data support a novel role for NARC-1/PCSK9 in CNS development, distinct from that in cholesterogenic organs such as liver.

Implication of the proprotein convertase NARC-1/PCSK9 in the development of the nervous system.

Neuronale apoptose r&eacute;gul&eacute;e convertase-1/proprot&eacute;ine convertase subtilisine-kexin ressemblant-9 (NARC-1/PCSK9) est un proprot&eacute;ine convertase r&eacute;cemment d&eacute;crite pour jouer un r&ocirc;le important dans l&#39;hom&eacute;ostasie du cholest&eacute;rol &agrave; travers la d&eacute;gradation accrue du r&eacute;cepteur LDL (LDLR) et, &eacute;ventuellement, dans le d&eacute;veloppement neural. Ici, nous avons &eacute;tudi&eacute; l&#39;implication potentielle de cette prot&eacute;inase dans le d&eacute;veloppement de la CNS utilisant des cellules embryonnaires pluripotentes P19 de souris et le poisson-z&egrave;bre comme mod&egrave;les. Temps parcours quantitatives RT-PCR analyses ont &eacute;t&eacute; effectu&eacute;es suite &agrave; l&#39;acide r&eacute;tino&iuml;que (RA)-induit la diff&eacute;renciation neuroectodermique de cellules P19. En cons&eacute;quence, les taux d&#39;ARNm de NARC-1/PCSK9 a culmin&eacute; &agrave; jour 2 de diff&eacute;renciation et est tomb&eacute; par la suite. En revanche, l&#39;expression des proprot&eacute;ines convertases subtilisine kexin isoenzyme 1/site 1 prot&eacute;ase et furine n&#39;est pas affect&eacute;e par la RA, tandis que celle du PC5/6 et PC2 a augment&eacute; au sein et/ou apr&egrave;s les 4 premiers jours de la p&eacute;riode de diff&eacute;renciation, respectivement. Ce mod&egrave;le n&#39;est pas affect&eacute; par la cholest&eacute;rog&eacute;n&egrave;se transcription factor st&eacute;rol r&eacute;glementaire &eacute;l&eacute;ment contraignant prot&eacute;ine-2, qui normalement haut-r&eacute;gule le taux de mRNA NARC-1/PCSK9 dans le foie. En outre, dans les cellules P19, traitement Antirhumatismal n&#39;affecte pas le niveau de prot&eacute;ine de l&#39;endog&egrave;ne LDLR. Ceci est en accord avec le mod&egrave;le d&#39;expression unique de NARC-1/PCSK9 chez le rongeur CNS, y compris le cervelet, o&ugrave; le g&egrave;ne LDLR n&#39;est pas nettement exprim&eacute;. Entier-Montez l&#39;hybridation in situ a r&eacute;v&eacute;l&eacute; que le patron d&#39;expression du poisson-z&egrave;bre NARC-1/PCSK9 est similaire &agrave; celle des souris dans le CNS et la p&eacute;riph&eacute;rie. Pr&eacute;cipitation sp&eacute;cifique du poisson-z&egrave;bre NARC-1/PCSK9 ARNm a entra&icirc;n&eacute; une d&eacute;sorganisation g&eacute;n&eacute;rale des neurones c&eacute;r&eacute;braux et perte des limites m&eacute;senc&eacute;phale rhombenc&eacute;phale, conduisant &agrave; une mortalit&eacute; embryonnaire &agrave; environ 96 h apr&egrave;s la f&eacute;condation. Ces donn&eacute;es appuient un nouveau r&ocirc;le pour la NARC-1/PCSK9 dans le d&eacute;veloppement de CNS, distinct de celle de cholest&eacute;rog&eacute;n&egrave;se organes comme le foie.

Implication de la proprot&eacute;ine convertase NARC-1/PCSK9 dans le d&eacute;veloppement du syst&egrave;me nerveux.

Neuronales Apoptosis reguliert Konvertase-1/proprotein-Konvertase Subtilisin-Kexin wie-9 (NARC-1/PCSK9) ist ein proprotein Konvertase k&uuml;rzlich beschrieben, um eine wichtige Rolle im Cholesterin-Hom&ouml;ostase durch verst&auml;rkten Abbau des Rezeptors low density Lipoprotein (LDLR) und m&ouml;glicherweise in neuronalen Entwicklung spielen. Hierin, untersuchten wir die m&ouml;gliche Beteiligung der diese Proteinase bei der Entwicklung des ZNS mit Maus embryonal pluripotenten P19 Zellen und der Zebrafisch als Modelle. Zeit Kurs quantitative RT-PCR Analysen durchgef&uuml;hrt wurden, nach Retins&auml;ure (RA)-induzierte Neuroektodermale Differenzierung P19-Zellen. Dementsprechend die mRNA-Niveaus der NARC-1/PCSK9 erreichte bei Tag 2 der Differenzierung und danach fiel. Im Gegensatz dazu war der Ausdruck der proprotein Convertases Subtilisin Kexin Isoenzym 1/Website 1 Protease und Furin unbeeinflusst von RA, w&auml;hrend der PC5/6 und PC2 innerhalb bzw. nach den ersten 4 Tagen der Periode Differenzierung zulegten. Dieses Muster wurde von der Cholesterogenic Transkription Faktor Sterin regulatorischen Element-Binding Protein-2, die normalerweise oben-NARC-1/PCSK9 mRNA-Stufen in Leber regelt, nicht betroffen. Dar&uuml;ber hinaus RA-Behandlung in P19-Zellen, die Protein-Ebene, der die endogene LDLR nicht betroffen. Dies stimmt &uuml;berein mit dem einzigartigen Ausdrucksmuster der NARC-1/PCSK9 in das Nagetier CNS, einschlie&szlig;lich das Kleinhirn, wo die LDLR nicht deutlich zum Ausdruck kommt. Ganz-Mount-in situ-Hybridisierung ergab, dass das Muster des Ausdrucks der Zebrafisch NARC-1/PCSK9 &auml;hnlich der Maus im CNS und Peripherie. Spezifische Niederschlag von Zebrafish NARC-1/PCSK9 mRNA f&uuml;hrte zu eine allgemeine Desorganisation zerebell&auml;re Neuronen und Verlust von Hinterhirn-Mittelhirn-Grenzen zu embryonalen Tod bei etwa 96 Stunden nach der Befruchtung. Diese Daten unterst&uuml;tzen eine neuartige Rolle f&uuml;r NARC-1/PCSK9 in CNS-Entwicklung, die sich von dem in Cholesterogenic Organen wie Leber.

Implikation der proprotein Konvertase NARC-1/PCSK9 bei der Entwicklung des Nervensystems.

Neurali convertasi convertasi-1/proprotein regolamentati apoptosi subtilisina-kexin simil-9 (NARC-1/PCSK9) &egrave; un proprotein convertasi recentemente descritto per giocare un ruolo importante nell&#39;omeostasi del colesterolo attraverso una maggiore degradazione del recettore della lipoproteina a bassa densit&agrave; (LDLR) e possibilmente nello sviluppo neurale. Nel presente documento, abbiamo studiato il potenziale coinvolgimento di questo proteinasi nello sviluppo del CNS utilizzando cellule embrionali pluripotenti P19 mouse e the zebrafish come modelli. Analisi di RT-PCR quantitative tempo corso sono state effettuate seguendo l&#39;acido retinoico (RA)-indotta neuroectodermica differenziazione delle cellule P19. Di conseguenza, i livelli di mRNA di NARC-1/PCSK9 raggiunse il giorno 2 di differenziazione e caddero in seguito. Al contrario, l&#39;espressione delle proteasi di proprotein convertasi subtilisina kexin isoenzimatici 1 sito 1 e Furin era influenzato da RA, mentre quello di PC5/6 e PC2 aumentata all&#39;interno e/o dopo i primi 4 giorni del periodo di differenziazione rispettivamente. Questo modello non &egrave; stato influenzato dal cholesterogenic trascrizione fattore sterolo normativo elemento vincolante proteine-2, che normalmente up-regola i livelli di mRNA PCSK9/NARC-1 nel fegato. Inoltre, in cellule P19, trattamento RA non influenzava il livello della proteina del LDLR endogeno. Ci&ograve; concorda con il pattern di espressione unica di NARC-1/PCSK9 in roditore CNS, compreso il cervelletto, dove il LDLR non &egrave; espressa in modo significativo. Intero-monta l&#39;ibridazione in situ ha rivelato che il pattern di espressione di zebrafish NARC-1/PCSK9 &egrave; simile a quello del mouse sia nel CNS e periferia. Specifica atterramento di zebrafish NARC-1/PCSK9 mRNA ha provocato una disorganizzazione generale dei neuroni cerebellari e perdita di confini hindbrain-mesencefalo, portando a morte embrionale a circa 96 ore dopo la fecondazione. Questi dati supportano un ruolo romanzo per NARC-1/PCSK9 nello sviluppo di CNS, distinto da quello in cholesterogenic organi come il fegato.

Implicazione della convertasi proprotein NARC-1/PCSK9 nello sviluppo del sistema nervoso.

&#x795E;&#x7D4C;&#x30A2;&#x30DD;&#x30C8;&#x30FC;&#x30B7;&#x30B9;&#x898F;&#x5236;&#x306E;&#x8EE2;&#x63DB;&#x9175;&#x7D20; 1/&#x30D7;&#x30ED;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8EE2;&#x63DB;&#x9175;&#x7D20;&#x30B5;&#x30C1;&#x30E9;&#x30A4;&#x30B7;&#x30F3; kexin &#x69D8; 9 &#xFF08;&#x9EBB;&#x85AC;&#x306E;&#x58F2;&#x4EBA;-1/PCSK9) &#x30B3;&#x30EC;&#x30B9;&#x30C6;&#x30ED;&#x30FC;&#x30EB;&#x6052;&#x5E38;&#x6027;&#x4F4E;&#x6BD4;&#x91CD;&#x30EA;&#x30DD;&#x86CB;&#x767D;&#x53D7;&#x5BB9;&#x4F53; (LDLR) &#x306E;&#x5206;&#x89E3;&#x4FC3;&#x9032;&#x3092;&#x901A;&#x3058;&#x3066;&#x3001;&#x304A;&#x305D;&#x3089;&#x304F;&#x795E;&#x7D4C;&#x306E;&#x958B;&#x767A;&#x306B;&#x5927;&#x304D;&#x306A;&#x5F79;&#x5272;&#x3092;&#x679C;&#x305F;&#x3059;&#x306B;&#x6700;&#x8FD1;&#x8AAC;&#x660E;&#x30D7;&#x30ED;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8EE2;&#x63DB;&#x9175;&#x7D20;&#x3067;&#x3059;&#x3002;&#x672C;&#x660E;&#x7D30;&#x66F8;&#x3067;&#x3053;&#x306E;&#x30D7;&#x30ED;&#x30C6;&#x30A3;&#x30CA;&#x30FC;&#x30BC; &#x30DE;&#x30A6;&#x30B9; P19 &#x591A;&#x80FD;&#x6027;&#x5E79;&#x7D30;&#x80DE;&#x3068;&#x3001;&#x30BC;&#x30D6;&#x30E9;&#x30D5;&#x30A3;&#x30C3;&#x30B7;&#x30E5;&#x3092;&#x30E2;&#x30C7;&#x30EB;&#x306B;&#x3057;&#x305F;&#x4E2D;&#x67A2;&#x795E;&#x7D4C;&#x7CFB;&#x306E;&#x958B;&#x767A;&#x306E;&#x6F5C;&#x5728;&#x7684;&#x306A;&#x95A2;&#x4E0E;&#x3092;&#x691C;&#x8A0E;&#x3057;&#x307E;&#x3057;&#x305F;&#x3002;&#x6642;&#x9593;&#x30B3;&#x30FC;&#x30B9;&#x5B9A;&#x91CF;&#x7684; RT-PCR &#x89E3;&#x6790;&#x306F;&#x3001;&#x30EC;&#x30C1;&#x30CE;&#x30A4;&#x30F3;&#x9178; (RA)&#x3001;&#x6B21;&#x3092;&#x884C;&#x3063;&#x305F;-&#x795E;&#x7D4C;&#x5916;&#x80DA;&#x8449;&#x6027; P19 &#x7D30;&#x80DE;&#x306E;&#x5206;&#x5316;&#x8A98;&#x5C0E;&#x3055;&#x308C;&#x307E;&#x3059;&#x3002;&#x3057;&#x305F;&#x304C;&#x3063;&#x3066;&#x3001;&#x9EBB;&#x85AC;&#x306E;&#x58F2;&#x4EBA;-1/PCSK9 &#x306E; mRNA &#x306E;&#x30EC;&#x30D9;&#x30EB;&#x306F;&#x5206;&#x5316;&#x3067; 2 &#x65E5;&#x76EE;&#x3092;&#x30D4;&#x30FC;&#x30AF;&#x306B;&#x3001;&#x305D;&#x306E;&#x5F8C;&#x843D;&#x3061;&#x305F;&#x3002;&#x5BFE;&#x7167;&#x7684;&#x306B;&#x3001;PC5/6&#x3001;PC2 &#x306E;&#x5185;&#x3084;&#x5206;&#x5316;&#x671F;&#x9593;&#x306E;&#x6700;&#x521D;&#x306E; 4 &#x65E5;&#x9593;&#x5F8C;&#x305D;&#x308C;&#x305E;&#x308C;&#x5897;&#x52A0;&#x3057;&#x305F;&#x306B;&#x5BFE;&#x3057;&#x30D7;&#x30ED;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x304A;&#x30B5;&#x30C1;&#x30E9;&#x30A4;&#x30B7;&#x30F3; kexin &#x30A2;&#x30A4;&#x30BD;&#x30B6;&#x30A4;&#x30E0; 1/&#x30B5;&#x30A4;&#x30C8; 1 &#x30D7;&#x30ED;&#x30C6;&#x30A2;&#x30FC;&#x30BC;&#x3068;&#x98A8;&#x9234;&#x306E;&#x767A;&#x73FE;&#x306F; RA &#x306B;&#x3088;&#x3063;&#x3066;&#x5F71;&#x97FF;&#x3055;&#x308C;&#x306A;&#x304B;&#x3063;&#x305F;&#x3002;&#x3053;&#x306E;&#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x306F; cholesterogenic &#x8EE2;&#x5199;&#x56E0;&#x5B50;&#x30B9;&#x30C6;&#x30ED;&#x30FC;&#x30EB;&#x898F;&#x5236;&#x8981;&#x7D20;&#x30D0;&#x30A4;&#x30F3;&#x30C7;&#x30A3;&#x30F3;&#x30B0; &#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA; - &#x901A;&#x5E38;&#x30A2;&#x30C3;&#x30D7;&#x5236;&#x5FA1; - &#x809D;&#x81D3;&#x3067;&#x306E;&#x9EBB;&#x85AC;&#x306E;&#x58F2;&#x4EBA;-1/PCSK9 mRNA &#x306E;&#x30EC;&#x30D9;&#x30EB; 2&#x3001;&#x306B;&#x3088;&#x3063;&#x3066;&#x5F71;&#x97FF;&#x3055;&#x308C;&#x306A;&#x304B;&#x3063;&#x305F;&#x3002;&#x3055;&#x3089;&#x306B;&#x3001;P19 &#x7D30;&#x80DE;&#x306B; RA &#x6CBB;&#x7642;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x30EC;&#x30D9;&#x30EB;&#x306E;&#x5185;&#x56E0;&#x6027;&#x306E; LDLR &#x5F71;&#x97FF;&#x3092;&#x53D7;&#x3051;&#x306A;&#x304B;&#x3063;&#x305F;.&#x3053;&#x308C;&#x306F;&#x3001;&#x9EBB;&#x85AC;&#x306E;&#x58F2;&#x4EBA;-1/PCSK9 &#x306E;&#x30E6;&#x30CB;&#x30FC;&#x30AF;&#x306A;&#x8868;&#x73FE;&#x306E;&#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x3067;&#x3001;&#x9F67;&#x6B6F;&#x985E;&#x3001;LDLR &#x5927;&#x5E45;&#x306B;&#x8868;&#x73FE;&#x3055;&#x308C;&#x3066;&#x3044;&#x306A;&#x3044;&#x5C0F;&#x8133;&#x3092;&#x542B;&#x3080;&#x4E2D;&#x67A2;&#x795E;&#x7D4C;&#x7CFB;&#x3001;&#x540C;&#x610F;&#x3057;&#x307E;&#x3059;&#x3002;&#x5168;&#x4F53;&#x30DE;&#x30A6;&#x30F3;&#x30C8; in situ &#x30CF;&#x30A4;&#x30D6;&#x30EA;&#x30C0;&#x30A4;&#x30BC;&#x30FC;&#x30B7;&#x30E7;&#x30F3; &#x30BC;&#x30D6;&#x30E9;&#x30D5;&#x30A3;&#x30C3;&#x30B7;&#x30E5;&#x306E;&#x767A;&#x73FE;&#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x304C;&#x9EBB;&#x85AC;&#x306E;&#x58F2;&#x4EBA;-1/PCSK9 &#x30DE;&#x30A6;&#x30B9;&#x4E2D;&#x67A2;&#x795E;&#x7D4C;&#x7CFB;&#x3068;&#x672B;&#x68A2;&#x306E;&#x4E21;&#x65B9;&#x306E;&#x3068;&#x540C;&#x69D8;&#x3067;&#x3042;&#x308B;&#x3053;&#x3068;&#x3092;&#x660E;&#x3089;&#x304B;&#x306B;&#x3057;&#x307E;&#x3057;&#x305F;&#x3002;&#x7279;&#x5B9A;&#x306E;&#x30CE;&#x30C3;&#x30AF;&#x30C0;&#x30A6;&#x30F3; &#x30BC;&#x30D6;&#x30E9;&#x30D5;&#x30A3;&#x30C3;&#x30B7;&#x30E5;&#x9EBB;&#x85AC;&#x306E;&#x58F2;&#x4EBA;-1/PCSK9 mRNA &#x306E;&#x5C0F;&#x8133;&#x795E;&#x7D4C;&#x306E;&#x4E00;&#x822C;&#x7684;&#x306A;&#x7121;&#x79E9;&#x5E8F;&#x3068;&#x83F1;&#x8133;&#x4E2D;&#x8133;&#x306E;&#x5883;&#x754C;&#x306F;&#x3001;&#x53D7;&#x7CBE;&#x5F8C;&#x7D04; 96 h &#x3067;&#x80DA;&#x306E;&#x6B7B;&#x306B;&#x3064;&#x306A;&#x304C;&#x308B;&#x306E;&#x640D;&#x5931;&#x3092;&#x3082;&#x305F;&#x3089;&#x3057;&#x305F;&#x3002;&#x3053;&#x308C;&#x3089;&#x306E;&#x30C7;&#x30FC;&#x30BF; CNS &#x958B;&#x767A;&#x3001;&#x809D;&#x81D3;&#x306A;&#x3069;&#x306E; cholesterogenic &#x306E;&#x81D3;&#x5668;&#x306B;&#x304B;&#x3089;&#x660E;&#x77AD;&#x306A;&#x9EBB;&#x85AC;&#x306E;&#x58F2;&#x4EBA;-1/PCSK9 &#x306E;&#x65B0;&#x898F;&#x5F79;&#x5272;&#x3092;&#x30B5;&#x30DD;&#x30FC;&#x30C8;&#x3057;&#x307E;&#x3059;&#x3002;

&#x795E;&#x7D4C;&#x7CFB;&#x306E;&#x767A;&#x9054;&#x306B;&#x304A;&#x3051;&#x308B;&#x30D7;&#x30ED;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8EE2;&#x63DB;&#x9175;&#x7D20;&#x9EBB;&#x85AC;&#x306E;&#x58F2;&#x4EBA;-1/PCSK9 &#x306E;&#x542B;&#x610F;&#x3002;

&#xC2E0;&#xACBD; apoptosis &#xADDC;&#xC81C; convertase-1/proprotein convertase subtilisin kexin &#xAC19;&#xC740;-9 (&#xB9C8;&#xC57D; &#xB2E8;&#xC18D; &#xBC18;-1/PCSK9)&#xB294; &#xCD5C;&#xADFC; &#xC800;&#xBC00;&#xB3C4; &#xC9C0; &#xB2E8;&#xBC31;&#xC9C8; &#xC218;&#xC6A9; &#xCCB4; (LDLR)&#xC758; &#xD5A5;&#xC0C1; &#xB41C; &#xC131;&#xB2A5; &#xC800;&#xD558;&#xB97C; &#xD1B5;&#xD574; &#xCF5C;&#xB808;&#xC2A4;&#xD14C;&#xB864; &#xD56D;&#xC0C1;&#xC131; &#xBC0F; &#xC2DC;&#xC2A4;&#xD15C;&#xC774; &#xC2E0;&#xACBD; &#xBC1C;&#xB2EC;&#xC5D0; &#xC911;&#xC694; &#xD55C; &#xC5ED;&#xD560;&#xC744; &#xC124;&#xBA85; &#xD558;&#xB294; proprotein convertase. &#xC5EC;&#xAE30;, &#xC6B0;&#xB9AC;&#xAC00; &#xB9C8;&#xC6B0;&#xC2A4; embryonal pluripotent P19 &#xC804;&#xC9C0;&#xC640; zebrafish &#xBAA8;&#xB378;&#xB85C; &#xC0AC;&#xC6A9; &#xD558; &#xC5EC; CNS&#xC758; &#xAC1C;&#xBC1C;&#xC5D0;&#xC11C;&#xC774; &#xAC00;&#xC218;&#xBD84;&#xD574;&#xC758; &#xC7A0;&#xC7AC;&#xC801;&#xC778; &#xAD00;&#xB828; &#xC870;&#xC0AC;. &#xC2DC;&#xAC04; &#xCF54;&#xC2A4; &#xC591;&#xC801; RT-PCR &#xBD84;&#xC11D; retinoic &#xC0B0; (RA)&#xC5D0; &#xB530;&#xB77C; &#xC218;&#xD589; &#xB41C;-neuroectodermal P19 &#xC138;&#xD3EC; &#xBD84;&#xD654;&#xB97C; &#xC720;&#xB3C4; &#xD569;&#xB2C8;&#xB2E4;. &#xC774; &#xB530;&#xB77C; &#xB9C8;&#xC57D;-1/PCSK9 mRNA &#xC218;&#xC900;&#xC758; &#xCC28;&#xBCC4;&#xD654;&#xC758; &#xB0A0; 2&#xC5D0; &#xB9CC;&#xC871; &#xD558; &#xACE0; &#xADF8; &#xD6C4; &#xB5A8;&#xC5B4;. &#xBC18;&#xBA74;, PC5/6&#xACFC; p c 2&#xC758; &#xAC01;&#xAC01; &#xB0B4;&#xC758; &#xBC0F; &#xCC28;&#xBCC4;&#xD654; &#xAE30;&#xAC04;&#xC758; &#xCCAB; 4 &#xC77C; &#xD6C4;&#xC5D0; &#xC99D;&#xAC00; &#xD558;&#xB294; &#xBC18;&#xBA74; proprotein convertases subtilisin kexin isozyme 1/&#xC0AC;&#xC774;&#xD2B8; 1 protease&#xC640; Furin&#xC758; &#xC2DD; RA, &#xC601;&#xD5A5;&#xC744; &#xC544;&#xB2C8;&#xC5C8;&#xB2E4;. &#xC774; &#xD328;&#xD134;&#xC740; cholesterogenic &#xC804;&#xC0AC; &#xC778;&#xC790; sterol &#xADDC;&#xC81C; &#xC694;&#xC18C; &#xBC14;&#xC778;&#xB529; &#xB2E8;&#xBC31;&#xC9C8;-2, &#xCD5C;&#xB300; &#xC77C;&#xBC18;&#xC801;&#xC73C;&#xB85C; &#xC870;&#xC808; &#xAC04;&#xC5D0;&#xC11C; &#xB9C8;&#xC57D;-1/PCSK9 mRNA &#xC218;&#xC900; &#xC601;&#xD5A5;&#xC744; &#xBC1B;&#xC9C0; &#xC54A;&#xC2B5;&#xB2C8;&#xB2E4;. &#xB610;&#xD55C;, P19 &#xC140;&#xC5D0;&#xC11C; RA &#xCE58;&#xB8CC; &#xBBF8;&#xCE58;&#xC9C0; &#xC54A; &#xC558; &#xC5B4; &#xB0B4; &#xC0DD; LDLR &#xB2E8;&#xBC31;&#xC9C8; &#xC218;&#xC900;. &#xC774; &#xC124;&#xCE58;&#xB958;&#xB294; LDLR&#xC740; &#xD06C;&#xAC8C; &#xD45C;&#xD604; &#xB418;&#xC9C0; &#xC18C; &#xB1CC;&#xB97C; &#xD3EC;&#xD568; &#xD558; &#xC5EC; CNS&#xC5D0; &#xB9C8;&#xC57D; &#xB2E8;&#xC18D; &#xBC18;-1/pcsk9&#xC758; &#xB3C5;&#xD2B9;&#xD55C; &#xC2DD; &#xD328;&#xD134;&#xC5D0; &#xB3D9;&#xC758; &#xD55C;&#xB2E4;. &#xC804;&#xCCB4; &#xB9C8;&#xC6B4;&#xD2B8; &#xC81C;&#xC790;&#xB9AC; &#xAD50; &#xC7A1; &#xC774;&#xB77C;&#xACE0; &#xBC1D;&#xD614;&#xB2E4; zebrafish&#xC758; &#xD45C;&#xD604; &#xD328;&#xD134; &#xB9C8;&#xC57D;-1/PCSK9 &#xB9C8;&#xC6B0;&#xC2A4; CNS&#xC640; &#xC8FC;&#xBCC0;&#xC5D0;&#xC11C; &#xBAA8;&#xB450; &#xBE44;&#xC2B7;&#xD569;&#xB2C8;&#xB2E4;. Zebrafish &#xB9C8;&#xC57D;-1/PCSK9 Mrna&#xC758; &#xD2B9;&#xC815; &#xCD5C;&#xC800;&#xC758; &#xC18C; &#xB1CC; &#xC2E0;&#xACBD; &#xC138;&#xD3EC;&#xC758; &#xC77C;&#xBC18;&#xC801;&#xC778; &#xD574;&#xCCB4; &#xD558; &#xACE0; &#xC218;&#xC815; &#xD6C4;&#xC5D0; &#xC57D; 96 h &#xBC30;&#xC544; &#xC0AC;&#xB9DD;&#xC5D0; &#xC774;&#xB974;&#xB294; hindbrain midbrain &#xACBD;&#xACC4;&#xC758; &#xC190;&#xC2E4;&#xC744; &#xCD08;&#xB798; &#xD588;&#xB2E4;. &#xC774;&#xB7EC;&#xD55C; &#xB370;&#xC774;&#xD130; CNS &#xAC1C;&#xBC1C;, &#xAC04; &#xB4F1; &#xC7A5;&#xAE30; cholesterogenic&#xB294; &#xB2E4;&#xB985;&#xB2C8;&#xB2E4; &#xB9C8;&#xC57D;-1/pcsk9&#xC5D0; &#xB300; &#xD55C; &#xC0C8;&#xB85C;&#xC6B4; &#xC5ED;&#xD560;&#xC744; &#xC9C0; &#xC6D0;&#xD558;&#xB294;.

&#xC2E0;&#xACBD; &#xC2DC;&#xC2A4;&#xD15C;&#xC758; &#xAC1C;&#xBC1C;&#xC5D0;&#xC11C; proprotein convertase &#xB9C8;&#xC57D;-1/pcsk9&#xC758; &#xC758;&#xBBF8;.

Convertasa convertasa-1/proprote&iacute;na regulados por apoptosis neuronal subtilisin-kexin como-9 (NARC-1/PCSK9) es una convertasa proprote&iacute;na descrito recientemente para desempe&ntilde;ar un papel importante en la homeostasis de colesterol a trav&eacute;s de mayor degradaci&oacute;n del receptor de la lipoprote&iacute;na de baja densidad (LDLR) y posiblemente en el desarrollo neuronal. Aqu&iacute;, se investig&oacute; la posible participaci&oacute;n de esta proteinasa en el desarrollo del SNC con c&eacute;lulas embrionarias pluripotenciales P19 de rat&oacute;n y pez cebra como modelo. Se realizaron an&aacute;lisis de RT-PCR cuantitativos tiempo curso siguiendo el &aacute;cido retinoico (RA)-inducida neuroectodermal diferenciaci&oacute;n de c&eacute;lulas P19. En consecuencia, los niveles de mRNA de NARC-1/PCSK9 alcanz&oacute; el d&iacute;a 2 de diferenciaci&oacute;n y cayeron despu&eacute;s de eso. En cambio, la expresi&oacute;n de la convertases proprote&iacute;na subtilisin kexin isoenzima 1/sitio 1 proteasa y Furin era inafectada por RA, mientras que la de PC5/6 y PC2 aument&oacute; dentro o despu&eacute;s de los primeros 4 d&iacute;as del per&iacute;odo de diferenciaci&oacute;n respectivamente. Este patr&oacute;n no fue afectado por la cholesterogenic transcripci&oacute;n factor esterol regulador elemento vinculante protein-2, que normalmente para arriba-regula los niveles de mRNA de NARC-1/PCSK9 en h&iacute;gado. Adem&aacute;s, en las c&eacute;lulas P19, RA tratamiento no afect&oacute; el nivel de prote&iacute;na de los LDLR end&oacute;geno. Esto coincide con el patr&oacute;n de expresi&oacute;n &uacute;nica de NARC-1/PCSK9 en el roedor CNS, incluyendo el cerebelo, donde los LDLR significativamente no se expresa. Hibridaci&oacute;n in situ de Whole-mount revel&oacute; que el patr&oacute;n de expresi&oacute;n del pez cebra NARC-1/PCSK9 es similar a la del rat&oacute;n en el CNS y periferia. Espec&iacute;fica ca&iacute;da del pez cebra NARC-1/PCSK9 mRNA dio lugar a una desorganizaci&oacute;n general de neuronas cerebelosas y p&eacute;rdida de l&iacute;mites del cerebelo-mesenc&eacute;falo, llevando a la muerte embrionaria en aproximadamente 96 h despu&eacute;s de la fertilizaci&oacute;n. Estos datos apoyan un papel nuevo para NARC-1/PCSK9 en el desarrollo de la CNS, distinto de la en &oacute;rganos de cholesterogenic como el h&iacute;gado.

Implicaci&oacute;n de la convertasa proprote&iacute;na NARC-1/PCSK9 en el desarrollo del sistema nervioso.

Lys-gamma3-MSH is a melanocortin peptide derived from the C-terminal of the 16 kDa fragment of POMC. The physiological role of Lys-gamma3-MSH is unclear, although it has previously been shown that, although not directly steroidogenic, it can act to potentiate the steroidogenic response of adrenal cortical cells to ACTH. This synergistic effect appears to be correlated with an ability to increase the activity of hormone sensitive lipase (HSL) and therefore the rate of cholesterol ester hydrolysis. Ligand binding studies have suggested that high-affinity binding sites for Lys-gamma3-MSH exist in the adrenal gland and a number of other rat tissues that express HSL, including adipose, skeletal muscle and testes. To investigate the hypothesis that Lys-gamma3-MSH may play a wider role in cholesterol and lipid metabolism, we tested the effect of Lys-gamma3-MSH on lipolysis, an HSL-mediated process, in 3T3-L1 adipocytes. In comparison with other melanocortin peptides, Lys-gamma3-MSH was found to be a potent stimulator of lipolysis. It was also able to phosphorylate HSL at key serine residues and stimulate the hyperphosphorylation of perilipin A. The receptor through which the lipolytic actions of Lys-gamma3-MSH are being mediated is not clear. Attempts to characterise this receptor suggest that either the pharmacology of the melanocortin receptor 5 in 3T3-L1 adipocytes is different from that described when expressed in heterologous systems or the possibility that a further, as yet uncharacterised, receptor exists.

Evidence of a possible role for Lys-gamma3-MSH in the regulation of adipocyte function.

Progranulin is a secreted protein with important functions in several physiological and pathological processes, such as embryonic development, host defense, and wound repair. Autosomal dominant mutations in the progranulin gene cause frontotemporal dementia, while overexpression of progranulin promotes the invasive progression of a range of tumors, including those of the breast and the brain. Structurally, progranulin consists of seven-and-a-half tandem repeats of the granulin/epithelin module (GEM), several of which have been isolated as discrete 6-kDa GEM peptides. We have expressed all seven human GEMs using recombinant DNA in Escherichia coli. High-resolution NMR showed that only the three GEMs, hGrnA, hGrnC, and hGrnF, contain relatively well-defined three-dimensional structures in solution, while others are mainly mixtures of poorly structured disulfide isomers. The three-dimensional structures of hGrnA, hGrnC, and hGrnF contain a stable stack of two beta-hairpins in their N-terminal subdomains, but showed a more flexible C-terminal subdomain. Interestingly, of the well-structured GEMs, hGrnA demonstrated potent growth inhibition of a breast cancer cell line, while hGrnF was stimulatory. Poorly folded peptides were either weakly inhibitory or without activity. The functionally active and structurally well-characterized human hGrnA offers a unique opportunity for detailed structure-function studies of these important GEM proteins as novel members of mammalian growth factors.

Structure dissection of human progranulin identifies well-folded granulin/epithelin modules with unique functional activities.

The growth factor progranulin (PGRN) regulates cell division, survival, and migration. PGRN is an extracellular glycoprotein bearing multiple copies of the cysteine-rich granulin motif. With PGRN family members in plants and slime mold, it represents one of the most ancient of the extracellular regulatory proteins still extant in modern animals. PRGN has multiple biological roles. It contributes to the regulation of early embryogenesis, to adult tissue repair and inflammation. Elevated PGRN levels often occur in cancers, and PGRN immunotherapy inhibits the growth of hepatic cancer xenografts in mice. Recent studies have demonstrated roles for PGRN in neurobiology. An autosomal dominant mutation in GRN, the gene for PGRN, leads to neuronal atrophy in the frontal and temporal lobes, resulting in the disease frontotemporal lobar dementia. In this review we will discuss current knowledge of the multifaceted biology of PGRN.

The granulin gene family: from cancer to dementia.

Progranulin is a secreted high molecular weight growth factor bearing seven and one half copies of the cysteine-rich granulin-epithelin motif. While inappropriate over-expression of the progranulin gene has been associated with many cancers, haploinsufficiency leads to atrophy of the frontotemporal lobes and development of a form of dementia (frontotemporal lobar degeneration with ubiquitin positive inclusions, FTLD-U) associated with the formation of ubiquitinated inclusions. Recent reports indicate that progranulin has neurotrophic effects, which, if confirmed would make progranulin the only neuroprotective growth factor that has been associated genetically with a neurological disease in humans. Preliminary studies indicated high progranulin gene expression in spinal cord motor neurons. However, it is uncertain what the role of Progranulin is in normal or diseased motor neuron function. We have investigated progranulin gene expression and subcellular localization in cultured mouse embryonic motor neurons and examined the effect of progranulin over-expression and knockdown in the NSC-34 immortalized motor neuron cell line upon proliferation and survival.

Progranulin is expressed within motor neurons and promotes neuronal cell survival.

Pro-protein convertase &#x67AF;&#x8349;/&#x79D1;&#x4FE1; 5 (PC5&#xFF0C;&#x4E5F;&#x79F0;&#x4E3A; PC6&#xFF09; &#x662F;&#x7275;&#x8FDE;&#x5230;&#x5176;&#x529F;&#x80FD;&#x6D3B;&#x8DC3;&#x7684;&#x884D;&#x751F;&#x5DE5;&#x5177;&#x7684;&#x6F5C;&#x5728;&#x524D;&#x4F53;&#x86CB;&#x767D;&#x6210;&#x719F;&#x7684;&#x4E1D;&#x6C28;&#x9178;&#x86CB;&#x767D;&#x9176;&#x7684;&#x67AF;&#x8349;&#x6746;&#x83CC;&#x5BB6;&#x65CF;&#x7684;&#x6210;&#x5458;&#x3002;&#x82E5;&#x8981;&#x8C03;&#x67E5;&#x7684;&#x804C;&#x80FD;&#x89D2;&#x8272;&#xFF0C;&#x6211;&#x4EEC;&#x6709;&#x514B;&#x9686;&#x7684; cDNA &#x5E8F;&#x5217;&#x7F16;&#x7801;&#x4E24;&#x4E2A;&#x5019;&#x9009;&#x4EBA;&#x6591;&#x9A6C;&#x9C7C; PC5 convertases &#xFF08;&#x6307;&#x5B9A;&#x4E3A; PCSK5.1 &#x548C; PCSK5.2&#xFF09; co-orthologous &#x5230;&#x5355;&#x4E2A; PC5 &#x7F16;&#x7801;&#x57FA;&#x56E0; (PCSK5) &#x5728;&#x54FA;&#x4E73;&#x52A8;&#x7269;&#x4E2D;&#x627E;&#x5230;&#x3002;&#x4E24;&#x8005;&#x90FD;&#x663E;&#x793A;&#x4EBA;&#x7C7B; PCSK5 &#x57FA;&#x56E0; syntenic &#x51FD;&#x6388;&#x3002;&#x6574;&#x4F53;&#x57FA;&#x56E0;&#x7ED3;&#x6784;&#x5DF2;&#x8DE8;&#x7269;&#x79CD;&#x5DF2;&#x4FDD;&#x5B88;&#x3002;&#x867D;&#x7136; PC5.1 mRNA &#x8868;&#x8FBE;&#x662F;&#x975E;&#x5E38;&#x79BB;&#x6563;&#x8033;&#x56CA;&#x6CE1;&#x548C;&#x4FA7;&#x7EBF; neuromasts &#x8303;&#x56F4;&#x5185;&#xFF0C;PC5.2 &#x6210;&#x7EE9;&#x5355;&#x66F4;&#x65E0;&#x5904;&#x4E0D;&#x5728;&#x4E0E;&#x7279;&#x5B9A;&#x7684;&#x672C;&#x5730;&#x5316;&#xFF0C;&#x5404;&#x673A;&#x6784;&#x5305;&#x62EC;&#x809D;&#x3001; &#x80A0;&#x3001;&#x3001; &#x8033;&#x6CE1;&#x5728;&#x4E2D;&#x67A2;&#x795E;&#x7ECF;&#x7CFB;&#x7EDF;&#x5185;&#x8868;&#x793A;&#x3002;&#x6591;&#x9A6C;&#x9C7C; PC5.1 &#x7F3A;&#x80DA;&#x80CE;&#x663E;&#x793A;&#x5F02;&#x5E38; neuromast &#x4FA7;&#x7EBF;&#x7CFB;&#x7EDF;&#x5185;&#x7684;&#x6C89;&#x79EF;&#x548C;&#x7F3A;&#x4E4F;&#x6B63;&#x5E38;&#x7684;&#x89E6;&#x6478;&#x53CD;&#x5E94;&#xFF0C;&#x4E0E;&#x4FA7;&#x7EBF;&#x5728;&#x5BF9;&#x7A7A;&#x95F4;&#x7684;&#x8BA4;&#x8BC6;&#x548C;&#x611F;&#x77E5;&#x73AF;&#x5883;&#x7684;&#x5DF2;&#x77E5;&#x611F;&#x5B98;&#x4F5C;&#x7528;&#x76F8;&#x4E00;&#x81F4;&#x3002;

&#x5206;&#x5B50;&#x514B;&#x9686;&#x548C;&#x80DA;&#x80CE;&#x8868;&#x8FBE;&#x7684;&#x6591;&#x9A6C;&#x9C7C; PCSK5 co orthologues&#xFF1A; &#x529F;&#x80FD;&#x8BC4;&#x4F30;&#x671F;&#x95F4;&#x4FA7;&#x7EBF;&#x53D1;&#x5C55;&#x3002;

Pro-protein convertase subtilisin/kexin 5 (PC5, also known as PC6) is a member of the subtilisin-like superfamily of serine proteases implicated in the maturation of latent precursor proteins into their functionally active derivatives. To investigate the functional roles, we have cloned the cDNA sequences encoding two candidate zebrafish PC5 convertases (designated as PCSK5.1 and PCSK5.2) co-orthologous to the single PC5 encoding gene (PCSK5) found in mammals. Both display syntenic correspondence to the human PCSK5 gene. Overall gene architecture has been conserved across species. While PC5.1 mRNA expression is very discrete within the otic vesicle and lateral line neuromasts, PC5.2 transcripts are more ubiquitously expressed within the central nervous system together with specific localization in various organs including liver, intestine, and otic vesicle. Zebrafish PC5.1-deficient embryos display abnormal neuromast deposition within the lateral line system and lack a normal touch response, consistent with the known sensory role that the lateral line plays in spatial awareness and sensing the environment.

Molecular cloning and embryonic expression of zebrafish PCSK5 co-orthologues: functional assessment during lateral line development.

Proprot&eacute;ines convertases subtilisine/kexin 5 (PC5, &eacute;galement connu sous le nom de PC6) est un membre de la superfamille de subtilisine des s&eacute;rines prot&eacute;ases impliqu&eacute;es dans la maturation des prot&eacute;ines pr&eacute;curseurs latente dans leurs d&eacute;riv&eacute;s fonctionnellement actifs. Afin d&#39;&eacute;tudier les r&ocirc;les fonctionnels, nous avons clon&eacute; les s&eacute;quences d&#39;ADNc codant des deux candidats zebrafish PC5 convertases (d&eacute;sign&eacute; comme PCSK5.1 et PCSK5.2) co-orthologue du PC5 unique codant le g&egrave;ne (PCSK5), chez les mammif&egrave;res. Les deux affichent synt&eacute;nie correspondance vers le g&egrave;ne humain de la PCSK5. Dans l&#39;ensemble, architecture de g&egrave;ne a &eacute;t&eacute; conserv&eacute; &agrave; travers les esp&egrave;ces. Tandis que l&#39;expression de l&#39;ARNm PC5.1 est tr&egrave;s discrete dans la v&eacute;sicule otique et neuromasts de la ligne lat&eacute;rale, les transcriptions PC5.2 sont exprim&eacute;es plus ubiquitaire dans le syst&egrave;me nerveux central ainsi que la localisation sp&eacute;cifique dans divers organes, y compris le foie, intestin et v&eacute;sicule otique. Des embryons de poisson-z&egrave;bre PC5.1 d&eacute;ficients en affichent des d&eacute;p&ocirc;ts anormaux de neuromast au sein du syst&egrave;me de la ligne lat&eacute;rale et n&#39;ont pas une r&eacute;ponse tactile normal, conforme au r&ocirc;le sensoriel connu que la ligne lat&eacute;rale joue dans la conscience spatiale et la t&eacute;l&eacute;d&eacute;tection de l&#39;environnement.

Clonage et expression embryon de poisson z&egrave;bre PCSK5 co-orthologues : &eacute;valuation fonctionnelle au cours du d&eacute;veloppement de la ligne lat&eacute;rale.

Pro-Protein-Konvertase Subtilisin/Kexin 5 (PC5, auch bekannt als PC6) ist ein Mitglied der Superfamilie Subtilisin-&auml;hnlichen Serinproteasen, die die Reifung der latente Vorl&auml;ufer-Proteine in ihre funktionell aktive Derivate in Verbindung gebracht. Um die Funktionsrollen zu untersuchen, haben wir die cDNA-Sequenzen, die Codierung von zwei Kandidaten Zebrafisch PC5 Convertases (bezeichnet als PCSK5.1 und PCSK5.2) co-Orthologous zu den einzelnen PC5 Codierung gen (PCSK5) gefunden bei S&auml;ugetieren geklont. Beide anzeigen syntenic Korrespondenz f&uuml;r das menschliche PCSK5-gen. Insgesamt hat gen Architektur Arten konserviert worden. W&auml;hrend PC5.1 mRNA Ausdruck sehr diskret innerhalb der otogenes Vesikel und Seitenlinienorgan Neuromasts ist, sind PC5.2 Abschriften mehr ubiquit&auml;r im Zentralnervensystem zusammen mit bestimmten Lokalisierung in verschiedenen Organen wie Leber, Darm und otogenes Vesikel ge&auml;u&szlig;ert. Zebrafish PC5.1-defizienten Embryonen zeigen abnorme Neuromast Ablagerung innerhalb des Systems Seitenlinienorgan und fehlen eine normalen Touch-Reaktion, mit der bekannten sensorische Funktion die Seitenlinie in r&auml;umliche Wahrnehmung und Erkennung der Umwelt vereinbar.

Molekulares Klonen und embryonalen Ausdruck von Zebrafish PCSK5 co-Orthologues: funktionale Bewertung w&auml;hrend der Seitenlinie Entwicklung.

Pro-protein convertasi subtilisina/kexin 5 (PC5, noto anche come PC6) &egrave; un membro della superfamiglia subtilisina-come delle proteasi serina implicati nella maturazione delle proteine precursore latente in loro derivati funzionalmente attivi. Per indagare i ruoli funzionali, noi abbiamo clonato le sequenze del cDNA che codifica due candidati zebrafish PC5 convertasi (indicato come PCSK5.1 e PCSK5.2) co-orthologous per il singolo PC5 codifica gene (PCSK5) trovato nei mammiferi. Entrambi visualizzare syntenic corrispondenza al gene umano PCSK5. Complessiva architettura gene conservato in tutta la specie. Mentre l&#39;espressione del mRNA PC5.1 &egrave; molto discreta all&#39;interno della vescicola otic e linea laterale neuromasts, PC5.2 trascrizioni sono pi&ugrave; ubiquitously espresso all&#39;interno del sistema nervoso centrale insieme a localizzazione specifica nei vari organi tra cui fegato, intestino e la vescicola otic. Embrioni di zebrafish PC5.1-carenti visualizzare neuromast anomala deposizione all&#39;interno del sistema della linea laterale e la mancanza di una risposta al tocco normale, coerenza con il ruolo sensoriale noto che la linea laterale gioca nella consapevolezza spaziale e l&#39;ambiente di rilevamento.

Clonazione molecolare e l&#39;espressione embrionale di Danio rerio PCSK5 co-orthologues: valutazione funzionale durante lo sviluppo della linea laterale.

Pro-protein &#x8EE2;&#x63DB;&#x9175;&#x7D20;&#x30B5;&#x30C1;&#x30E9;&#x30A4;&#x30B7;&#x30F3;/kexin 5 (PC5 PC6 &#x3068;&#x3082;&#x547C;&#x3070;&#x308C;&#x307E;&#x3059;) &#x306F;&#x3001;&#x30BB;&#x30EA;&#x30F3;&#x30D7;&#x30ED;&#x30C6;&#x30A2;&#x30FC;&#x30BC;&#x6F5C;&#x5728;&#x524D;&#x99C6;&#x4F53;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x306E;&#x6210;&#x719F;&#x306E;&#x6A5F;&#x80FD;&#x7684;&#x6D3B;&#x6027;&#x8A98;&#x5C0E;&#x4F53;&#x3078;&#x306E;&#x95A2;&#x4E0E;&#x306E;&#x30B5;&#x30C1;&#x30E9;&#x30A4;&#x30B7;&#x30F3;&#x69D8;&#x30B9;&#x30FC;&#x30D1;&#x30FC;&#x30D5;&#x30A1;&#x30DF;&#x30EA;&#x30FC;&#x306E;&#x30E1;&#x30F3;&#x30D0;&#x30FC;&#x3067;&#x3059;&#x3002;&#x6A5F;&#x80FD;&#x7684;&#x5F79;&#x5272;&#x3092;&#x8ABF;&#x67FB;&#x3059;&#x308B;&#x306B;&#x306F;&#x3001;2 &#x3064;&#x5019;&#x88DC;&#x30BC;&#x30D6;&#x30E9;&#x30D5;&#x30A3;&#x30C3;&#x30B7;&#x30E5; PC5 &#xFF08;PCSK5.1 &#x3068; PCSK5.2 &#x3068;&#x3057;&#x3066;&#x6307;&#x5B9A;&#xFF09; &#x304A; co-&#x54FA;&#x4E73;&#x985E;&#x3067;&#x767A;&#x898B;&#x907A;&#x4F1D;&#x5B50; (PCSK5) &#x306E;&#x30A8;&#x30F3;&#x30B3;&#x30FC;&#x30C7;&#x30A3;&#x30F3;&#x30B0;&#x306B;&#x307E;&#x305F;&#x304C;&#x308B;&#x30AA;&#x30EB;&#x30BD;&#x30ED;&#x30B0;&#x95A2;&#x4FC2;&#x5358;&#x4E00; PC5 &#x30A8;&#x30F3;&#x30B3;&#x30FC;&#x30C7;&#x30A3;&#x30F3;&#x30B0; cDNA &#x914D;&#x5217;&#x306E;&#x30AF;&#x30ED;&#x30FC;&#x30F3;&#x3092;&#x4F5C;&#x6210;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x4E21;&#x65B9;&#x30DD;&#x30B8;&#x30B7;&#x30E7;&#x30CA;&#x30EB;&#x30B7;&#x30F3;&#x30C6;&#x30CB;&#x30C3;&#x30AF;&#x5BFE;&#x5FDC; PCSK5 &#x907A;&#x4F1D;&#x5B50;&#x3092;&#x8868;&#x793A;&#x3057;&#x307E;&#x3059;&#x3002;&#x5168;&#x4F53;&#x7684;&#x306B;&#x7A2E;&#x3092;&#x6E21;&#x3063;&#x3066;&#x907A;&#x4F1D;&#x5B50;&#x30A2;&#x30FC;&#x30AD;&#x30C6;&#x30AF;&#x30C1;&#x30E3;&#x3092;&#x4FDD;&#x5B58;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;PC5.1 mRNA &#x306E;&#x767A;&#x73FE;&#x306F;&#x8033;&#x80DE;&#x3068;&#x5074;&#x7DDA;&#x30AA;&#x30A4;&#x30AB;&#x30EF;&#x5185;&#x975E;&#x5E38;&#x306B;&#x96E2;&#x6563;&#x7684;&#x3067;&#x3059;&#x304C;&#x3001;&#x4E2D;&#x67A2;&#x795E;&#x7D4C;&#x7CFB;&#x4E00;&#x7DD2;&#x306B;&#x7279;&#x5B9A;&#x306E;&#x30ED;&#x30FC;&#x30AB;&#x30EA;&#x30BC;&#x30FC;&#x30B7;&#x30E7;&#x30F3;&#x306E;&#x809D;&#x81D3;&#x3001;&#x8178;&#x3001;&#x8033;&#x80DE;&#x306A;&#x3069;&#x69D8;&#x3005; &#x306A;&#x81D3;&#x5668;&#x5185;&#x306E; PC5.2 &#x30C8;&#x30E9;&#x30F3; &#x30B9;&#x30AF;&#x30EA;&#x30D7;&#x30C8;&#x3088;&#x308A;&#x904D;&#x5728;&#x3067;&#x8868;&#x3055;&#x308C;&#x307E;&#x3059;&#x3002;PC5.1 &#x6B20;&#x640D;&#x30BC;&#x30D6;&#x30E9;&#x30D5;&#x30A3;&#x30C3;&#x30B7;&#x30E5;&#x80DA;&#x7570;&#x5E38; neuromast &#x84B8;&#x7740;&#x5074;&#x7DDA;&#x30B7;&#x30B9;&#x30C6;&#x30E0;&#x5185;&#x3067;&#x306E;&#x8868;&#x793A;&#x3057;&#x3001;&#x901A;&#x5E38;&#x30BF;&#x30C3;&#x30C1;&#x5FDC;&#x7B54;&#x3001;&#x5074;&#x7DDA;&#x7A7A;&#x9593;&#x610F;&#x8B58;&#x3068;&#x74B0;&#x5883;&#x3092;&#x611F;&#x77E5;&#x3067;&#x679C;&#x305F;&#x3057;&#x3066;&#x3044;&#x308B;&#x77E5;&#x3089;&#x308C;&#x3066;&#x3044;&#x308B;&#x611F;&#x899A;&#x306E;&#x5F79;&#x5272;&#x3068;&#x4E00;&#x8CAB;&#x6027;&#x3092;&#x6B20;&#x3044;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;

&#x5206;&#x5B50;&#x30AF;&#x30ED;&#x30FC;&#x30CB;&#x30F3;&#x30B0;&#x3068;&#x80DA;&#x6027;&#x767A;&#x73FE;&#x30BC;&#x30D6;&#x30E9;&#x30D5;&#x30A3;&#x30C3;&#x30B7;&#x30E5; PCSK5 co &#x7B2C;&#x30AA;&#x30FC;&#x30BD;&#x30ED;&#x30AC;&#x30B9;: &#x6A5F;&#x80FD;&#x7684;&#x30A2;&#x30BB;&#x30B9;&#x30E1;&#x30F3;&#x30C8;&#x5074;&#x7DDA;&#x958B;&#x767A;&#x4E2D;&#x3002;

Pro-protein convertase subtilisin/kexin 5 (PC5&#xB85C; &#xC54C;&#xB824;&#xC9C4; &#xB610;&#xD55C; PC6) &#xADF8;&#xB4E4;&#xC758; &#xAE30;&#xB2A5;&#xC801;&#xC73C;&#xB85C; &#xD65C;&#xC131; &#xC720;&#xB3C4;&#xCCB4;&#xC5D0; &#xC7A0;&#xC7AC; &#xB41C; &#xC804;&#xC870; &#xB2E8;&#xBC31;&#xC9C8;&#xC758; &#xC131;&#xC219;&#xC5D0; &#xC5F0;&#xB8E8; &#xB5A0;&#xB4E4;&#xACE0; &#xD504;&#xB85C; &#xD14C;&#xC544;&#xC758; subtilisin &#xAC19;&#xC740; superfamily&#xC758; &#xAD6C;&#xC131;&#xC6D0;&#xC785;&#xB2C8;&#xB2E4;. &#xC6B0;&#xB9AC; &#xB450; &#xD6C4;&#xBCF4; zebrafish PC5 (PCSK5.1 &#xBC0F; PCSK5.2&#xB85C; &#xC9C0;&#xC815;) convertases &#xACF5;&#xB3D9;-orthologous &#xB2E8;&#xC77C; pc5&#xC5D0; &#xC720;&#xC804;&#xC790; (PCSK5) &#xD3EC;&#xC720;&#xB958;&#xC5D0; &#xC778;&#xCF54;&#xB529;&#xC744; &#xC778;&#xCF54;&#xB529; cDNA &#xC2DC;&#xD000;&#xC2A4; &#xBCF5;&#xC81C; &#xAE30;&#xB2A5; &#xC5ED;&#xD560;&#xC744; &#xC870;&#xC0AC;. &#xB458; &#xB2E4; &#xC778;&#xAC04;&#xC758; PCSK5 &#xC720;&#xC804;&#xC790;&#xC5D0; syntenic &#xD1B5;&#xC2E0;&#xC744; &#xD45C;&#xC2DC;&#xD569;&#xB2C8;&#xB2E4;. &#xC804;&#xBC18;&#xC801;&#xC73C;&#xB85C; &#xC720;&#xC804;&#xC790; &#xC544;&#xD0A4;&#xD14D;&#xCC98; &#xC885;&#xC5D0; &#xAC78;&#xCCD0; &#xBCF4;&#xC874; &#xB418;&#xC5B4; &#xC788;&#xB2E4;. PC5.1 mRNA &#xC2DD;&#xC774; &#xB9E4;&#xC6B0; otic &#xAE30; &#xBC0F; &#xCE21;&#xBA74; &#xB77C;&#xC778; neuromasts &#xB0B4; &#xAC1C;&#xBCC4; &#xB3D9;&#xC548; PC5.2 &#xC131;&#xC801;&#xD45C;&#xB294; &#xB354; &#xD3B8; &#xAC04;, &#xCC3D; &#xC790;, otic &#xAE30; &#xB4F1; &#xB2E4;&#xC591; &#xD55C; &#xAE30;&#xAD00;&#xC5D0;&#xC11C; &#xD2B9;&#xC815; &#xC9C0;&#xC5ED;&#xD654;&#xC640; &#xC911;&#xC559; &#xC2E0; &#xACBD;&#xACC4; &#xB0B4;&#xC5D0;&#xC11C; &#xD45C;&#xC2DC; &#xB429;&#xB2C8;&#xB2E4;. PC5.1 &#xBD88;&#xCDA9;&#xBD84; &#xD55C; Zebrafish &#xD0DC;&#xC544; &#xCE21;&#xBA74; &#xB77C;&#xC778; &#xC2DC;&#xC2A4;&#xD15C; &#xB0B4;&#xC5D0;&#xC11C; &#xBE44;&#xC815;&#xC0C1;&#xC801;&#xC778; neuromast &#xC99D; &#xCC29;&#xC744; &#xD45C;&#xC2DC; &#xD558; &#xACE0; &#xC77C;&#xBC18; &#xD130;&#xCE58; &#xC751;&#xB2F5;, &#xC77C;&#xAD00;&#xC131; &#xCE21;&#xBA74; &#xB77C;&#xC778;&#xC740; &#xACF5;&#xAC04; &#xC778;&#xC2DD;&#xACFC; &#xD658;&#xACBD; &#xAC10;&#xC9C0;&#xC5D0; &#xC54C;&#xB824;&#xC9C4;&#xB41C; &#xAC10;&#xAC01; &#xC5ED;&#xD560;&#xC744; &#xBD80;&#xC871;.

&#xBD84;&#xC790; &#xD074;&#xB85C;&#xB2DD; &#xBC0F; zebrafish PCSK5 co orthologues &#xBC30;&#xC544; &#xD45C;&#xD604;&#xC758;: &#xCE21;&#xBA74; &#xB77C;&#xC778; &#xAC1C;&#xBC1C; &#xD558;&#xB294; &#xB3D9;&#xC548; &#xAE30;&#xB2A5; &#xD3C9;&#xAC00;.

Pro-Protein convertasa subtilisin/kexin 5 (PC5, PC6 tambi&eacute;n conocido como) es un miembro de la Superfamilia subtilisin-como de proteasas de serina implicado en la maduraci&oacute;n de prote&iacute;nas precursoras latente en sus derivados funcionalmente activos. Para investigar los papeles funcionales, hemos reproducido las secuencias de cDNA codificaci&oacute;n dos candidato pez cebra PC5 convertases (se&ntilde;alado como PCSK5.1 y PCSK5.2) co-orthologous a la sola PC5 codificaci&oacute;n gen (PCSK5) que se encuentra en los mam&iacute;feros. Ambas muestran correspondencia syntenic para el gen humano de PCSK5. En general ha sido conservada arquitectura gen&eacute;tica entre especies. Mientras que la expresi&oacute;n del mRNA PC5.1 es muy discreta dentro de la ves&iacute;cula &oacute;tica y neuromasts de la l&iacute;nea lateral, PC5.2 transcripciones m&aacute;s ubicuo se expresan dentro de sistema nervioso central junto con localizaci&oacute;n espec&iacute;fica en varios &oacute;rganos como el h&iacute;gado, el intestino y la ves&iacute;cula &oacute;tica. Embriones de pez cebra PC5.1 deficientes Mostrar la deposici&oacute;n anormal neuromast dentro del sistema de la l&iacute;nea lateral y carecen de una respuesta t&aacute;ctil normal, coherente con la funci&oacute;n sensorial conocida que la l&iacute;nea lateral desempe&ntilde;a en conciencia espacial y la percepci&oacute;n del medio ambiente.

Clonaci&oacute;n molecular y expresi&oacute;n embrionario del pez cebra PCSK5 co-orthologues: evaluaci&oacute;n funcional durante el desarrollo de la l&iacute;nea lateral.

Mutation of human GRN, the gene encoding the secreted glycoprotein progranulin, results in a form of frontotemporal lobar degeneration that is characterized by the presence of ubiquitinated inclusions containing phosphorylated and cleaved fragments of the transactivation response element DNA-binding protein-43. This has stimulated interest in understanding the role of progranulin in the central nervous system, and in particular, how this relates to neurodegeneration. Progranulin has many roles outside the brain, including regulation of cellular proliferation, survival, and migration, in cancer, including cancers of the brain, in wound repair, and inflammation. It often acts through the extracellular signal-regulated kinase and phopshatidylinositol-3-kinases pathways. The neurobiology of progranulin has followed a similar pattern with proposed roles for progranulin (PGRN) in the central nervous system as a neuroprotective agent and in neuroinflammation. Here we review the structure, biology, and mechanism of progranulin action. By understanding PGRN in a wider context, we may be better able to delineate its roles in the normal brain and in neurodegenerative disease.

Structure, function, and mechanism of progranulin; the brain and beyond.

The Zebrafish (Danio rerio) is a powerful and well-established tool used extensively for the study of early vertebrate development and as a model of human diseases. Zebrafish genes orthologous to their mammalian counterparts generally share conserved biological function. Protein knockdown or overexpression can be effectively achieved by microinjection of morpholino antisense oligonucleotides (MOs) or mRNA, respectively, into developing embryos at the one- to two-cell stage. Correlating gene expression patterns with the characterizing of phenotypes resulting from over- or underexpression can reveal the function of a particular protein. The microinjection technique is simple and results are reproducible. We defined the expression pattern of the proprotein convertase PCSK5 within the lateral line neuromasts and various organs including the liver, gut and otic vesicle by whole-mount in situ hybridization (ISH) and immunofluorescence (IF). MO-mediated knockdown of zebrafish PCSK5 expression generated embryos that display abnormal neuromast deposition within the lateral line system resulting in uncoordinated patterns of swimming.

Use of zebrafish and knockdown technology to define proprotein convertase activity.

The DNA/RNA binding proteins TAR DNA-binding protein 43 (TDP-43) and fused-in-sarcoma (FUS) are genetically linked to amyotrophic lateral sclerosis and frontotemporal lobar dementia, while the inappropriate cytoplasmic accumulations of TDP-43 and FUS are observed in a growing number of late-onset pathologies including spinocerebellar ataxia 3, Alzheimer's and Huntington's diseases (HD). To investigate if TDP-43 and FUS contribute to neurodegenerative phenotypes, we turned to a genetically accessible Caenorhabditis elegans model of polyglutamine toxicity. In C. elegans, we observe that genetic loss-of-function mutations for nematode orthologs of TDP-43 or FUS reduced behavioral defects and neurodegeneration caused by huntingtin exon-1 with expanded polyglutamines. Furthermore, using striatal cells from huntingtin knock-in mice we observed that small interfering ribonucleic acid (siRNA) against TDP-43 or FUS reduced cell death caused by mutant huntingtin. Moreover, we found that TDP-43 and the survival factor progranulin (PGRN) genetically interact to regulate polyglutamine toxicity in C. elegans and mammalian cells. Altogether our data point towards a conserved function for TDP-43 and FUS in promoting polyglutamine toxicity and that delivery of PGRN may have therapeutic benefits.

Article	Year
Solution structures of a 30-residue amino-terminal domain of the carp granulin-1 protein and its amino-terminally truncated 3-30 subfragment: implications for the conformational stability of... Proteins\| PubMed ID: 11870861	2002
The zebrafish progranulin gene family and antisense transcripts. BMC genomics\| PubMed ID: 16277664	2005
Implication of the proprotein convertase NARC-1/PCSK9 in the development of the nervous system. Journal of neurochemistry\| PubMed ID: 16893422	2006
Evidence of a possible role for Lys-gamma3-MSH in the regulation of adipocyte function. The Journal of endocrinology\| PubMed ID: 18180326	2008
Structure dissection of human progranulin identifies well-folded granulin/epithelin modules with unique functional activities. Protein science : a publication of the Protein Society\| PubMed ID: 18359860	2008
The granulin gene family: from cancer to dementia. BioEssays : news and reviews in molecular, cellular and developmental biology\| PubMed ID: 19795409	2009
Progranulin is expressed within motor neurons and promotes neuronal cell survival. BMC neuroscience\| PubMed ID: 19860916	2009
Molecular cloning and embryonic expression of zebrafish PCSK5 co-orthologues: functional assessment during lateral line development. Developmental dynamics : an official publication of the American Association of Anatomists\| PubMed ID: 20882679	2010
Structure, function, and mechanism of progranulin; the brain and beyond. Journal of molecular neuroscience : MN\| PubMed ID: 21691802	2011
Use of zebrafish and knockdown technology to define proprotein convertase activity. Methods in molecular biology (Clifton, N.J.)\| PubMed ID: 21805249	2011
Reduction of polyglutamine toxicity by TDP-43, FUS and progranulin in Huntington's disease models. Human molecular genetics\| PubMed ID: 23172908	2013

Hugh P. J. Bennett

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