reverse-genetics-mediated-recovery-of-infectious-murine-norovirus

Reverse Genetics Mediated Recovery of Infectious Murine Norovirus

Human noroviruses are responsible for most cases of human gastroenteritis (GE) worldwide and are recurrent problem in environments where close ...

Imperial College London 

Two features of viral quasispecies are reviewed: the presence of memory genomes as minority components of their mutant spectra, and viral extinction due to enhanced mutagenesis. Memory has been documented with several genetic markers of the important animal picornavirus foot-and-mouth disease virus (FMDV). The presence of memory genomes in viral quasispecies may accelerate their adaptive response whenever a selective constraint has already been experienced by a viral population during previous stages of its evolution. Enhanced mutagenesis has been shown to lead to losses of infectivity of a number of RNA viruses: poliovirus, vesicular stomatitis virus, human immunodeficiency virus type 1 and FMDV. These observations, based on the theoretical prediction of the existence of a copying error-threshold for maintenance of genetic information, may contribute to the development of a new antiviral strategy.

Emergence and selection of RNA virus variants: memory and extinction.

&#x5BA1;&#x67E5;&#x4E86;&#x75C5;&#x6BD2;&#x51C6;&#x79CD;&#x7684;&#x4E24;&#x4E2A;&#x7279;&#x70B9;&#xFF1A; &#x4F5C;&#x4E3A;&#x5176;&#x7A81;&#x53D8;&#x8C31;&#xFF0C;&#x548C;&#x589E;&#x5F3A;&#x8BF1;&#x53D8;&#x75C5;&#x6BD2;&#x6027;&#x540E;&#x7EE7;&#x4E4F;&#x4EBA;&#x7684;&#x5C11;&#x6570;&#x7EC4;&#x4EF6;&#x7684;&#x5185;&#x5B58;&#x57FA;&#x56E0;&#x7EC4;&#x5B58;&#x5728;&#x3002;&#x4E0E;&#x51E0;&#x4E2A;&#x9057;&#x4F20;&#x6807;&#x8BB0;&#x7684;&#x91CD;&#x8981;&#x52A8;&#x7269;&#x80A0;&#x9053;&#x75C5;&#x6BD2;&#x53E3;&#x75C5;&#x75C5;&#x6BD2; (&#x53E3;&#x8E44;&#x75AB;&#x75C5;&#x6BD2;) &#x8BB0;&#x8F7D;&#x4E86;&#x5185;&#x5B58;&#x3002;&#x6BCF;&#x5F53;&#x4E00;&#x4E2A;&#x9009;&#x62E9;&#x6027;&#x7684;&#x7EA6;&#x675F;&#x5DF2;&#x7ECF;&#x53D7;&#x5230;&#x75C5;&#x6BD2;&#x6027;&#x4EBA;&#x53E3;&#x5728;&#x524D;&#x4E00;&#x9636;&#x6BB5;&#x53CA;&#x5176;&#x6F14;&#x53D8;&#x7684;&#x8BB0;&#x5FC6;&#x57FA;&#x56E0;&#x7EC4;&#x4E2D;&#x75C5;&#x6BD2;&#x51C6;&#x79CD;&#x7684;&#x5B58;&#x5728;&#x53EF;&#x80FD;&#x4F1A;&#x52A0;&#x901F;&#x5176;&#x9002;&#x5E94;&#x6027;&#x53CD;&#x5E94;&#x3002;&#x5DF2;&#x8BC1;&#x660E;&#x589E;&#x5F3A;&#x7684;&#x7A81;&#x53D8;&#x5BFC;&#x81F4;&#x635F;&#x5931;&#x5927;&#x91CF;&#x7684; RNA &#x75C5;&#x6BD2;&#x7684;&#x4F20;&#x67D3;&#x6027;&#xFF1A; &#x810A;&#x9AD3;&#x7070;&#x8D28;&#x708E;&#x75C5;&#x6BD2;&#x3001; &#x6C34;&#x75B1;&#x6027;&#x53E3;&#x708E;&#x75C5;&#x6BD2;&#x3001; &#x4EBA;&#x7C7B;&#x514D;&#x75AB;&#x7F3A;&#x9677;&#x75C5;&#x6BD2; 1 &#x578B;&#x53E3;&#x8E44;&#x75AB;&#x75C5;&#x6BD2;&#x3002;&#x8FD9;&#x4E9B;&#x610F;&#x89C1;&#xFF0C;&#x57FA;&#x4E8E;&#x7406;&#x8BBA;&#x9884;&#x6D4B;&#x7684;&#x590D;&#x5236;&#x9519;&#x8BEF;&#x9608;&#x503C;&#xFF0C;&#x4EE5;&#x4FBF;&#x8FDB;&#x884C;&#x7EF4;&#x4FEE;&#x5DE5;&#x7A0B;&#x7684;&#x9057;&#x4F20;&#x4FE1;&#x606F;&#xFF0C;&#x5B58;&#x5728;&#x7684;&#x53EF;&#x80FD;&#x7684;&#x4E00;&#x79CD;&#x65B0;&#x7684;&#x6297;&#x75C5;&#x6BD2;&#x836F;&#x7269;&#x6218;&#x7565;&#x53D1;&#x5C55;&#x4F5C;&#x51FA;&#x8D21;&#x732E;&#x3002;

&#x51FA;&#x73B0;&#x548C; RNA &#x75C5;&#x6BD2;&#x53D8;&#x79CD;&#x7684;&#x9009;&#x62E9;&#xFF1A; &#x5185;&#x5B58;&#x548C;&#x706D;&#x7EDD;&#x3002;

Deux caract&eacute;ristiques des quasi-esp&egrave;ces virales sont pass&eacute;s en revue : la pr&eacute;sence des g&eacute;nomes de m&eacute;moire comme composants minoritaires de leurs spectres mutantes et extinction virale en raison de la mutagen&egrave;se am&eacute;lior&eacute;e. M&eacute;moire a &eacute;t&eacute; document&eacute; avec plusieurs marqueurs g&eacute;n&eacute;tiques du virus important picornavirus animaux pied &ndash; la maladie (fi&egrave;vre aphteuse). La pr&eacute;sence des g&eacute;nomes de m&eacute;moire en quasi-esp&egrave;ces virales peut-&ecirc;tre acc&eacute;l&eacute;rer leur r&eacute;ponse adaptative chaque fois qu&#39;une contrainte s&eacute;lective a d&eacute;j&agrave; &eacute;t&eacute; v&eacute;cue par une population virale au cours des pr&eacute;c&eacute;dentes &eacute;tapes de son &eacute;volution. Mutagen&egrave;se am&eacute;lior&eacute; a &eacute;t&eacute; d&eacute;montr&eacute; que conduire &agrave; des pertes d&#39;infectiosit&eacute; d&#39;un certain nombre de virus &agrave; ARN: poliovirus, virus de la stomatite v&eacute;siculaire, virus de l&#39;immunod&eacute;ficience humaine de type 1 et fi&egrave;vre aphteuse. Ces observations, fond&eacute;es sur la pr&eacute;diction th&eacute;orique de l&#39;existence d&#39;un seuil d&#39;erreur copie pour l&#39;entretien de l&#39;information g&eacute;n&eacute;tique, peuvent contribuer &agrave; l&#39;&eacute;laboration d&#39;une nouvelle strat&eacute;gie antivirale.

&Eacute;mergence et s&eacute;lection de variantes de virus &agrave; ARN : m&eacute;moire et extinction.

Entstehung und Selektion von RNA-Virus-Varianten: Ged&#xE4;chtnis und vom Aussterben bedroht.

Due caratteristiche delle quasispecie virali sono rivisti: la presenza di genomi di memoria come componenti di minoranza dei loro spettri mutanti e virale estinzione a causa della mutagenesi rafforzata. Memoria &egrave; stato documentato con diversi marcatori genetici del virus picornavirus animale importante foot and mouth disease (FMDV). La presenza di genomi di memoria nella quasispecie virale pu&ograve; accelerare la loro risposta adattativa ogni volta che un vincolo selettivo &egrave; gi&agrave; stato vissuto da una popolazione virale nelle fasi precedenti della sua evoluzione. Migliorata la mutagenesi ha dimostrata di portare a perdite di infettivit&agrave; di un certo numero di virus a RNA: poliovirus, virus della stomatite vescicolare, virus dell&#39;immunodeficienza umana di tipo 1 e FMDV. Queste osservazioni, basata sulla previsione teorica dell&#39;esistenza di una soglia di errore copia per la manutenzione delle informazioni genetiche, possono contribuire allo sviluppo di una nuova strategia antivirale.

Emersione e selezione delle varianti di virus a RNA: memoria ed estinzione.

&#x30A6;&#x30A4;&#x30EB;&#x30B9;&#x306E;&#x6E96;&#x7A2E;&#x306E; 2 &#x3064;&#x306E;&#x6A5F;&#x80FD;&#x3092;&#x30EC;&#x30D3;&#x30E5;&#x30FC;: &#x30E1;&#x30E2;&#x30EA; &#x30B2;&#x30CE;&#x30E0;&#x306E;&#x5B58;&#x5728;&#x305D;&#x306E;&#x30A6;&#x30A4;&#x30EB;&#x30B9;&#x6D88;&#x708E;&#x5F37;&#x5316;&#x3055;&#x308C;&#x305F;&#x7A81;&#x7136;&#x5909;&#x7570;&#x8A98;&#x767A;&#x5909;&#x7570;&#x30B9;&#x30DA;&#x30AF;&#x30C8;&#x30EB;&#x306E;&#x5C11;&#x6570;&#x306E;&#x30B3;&#x30F3;&#x30DD;&#x30FC;&#x30CD;&#x30F3;&#x30C8;&#x3068;&#x3057;&#x3066;&#x3002;&#x30E1;&#x30E2;&#x30EA;&#x3068;&#x3044;&#x304F;&#x3064;&#x304B;&#x306E;&#x907A;&#x4F1D;&#x7684;&#x30DE;&#x30FC;&#x30AB;&#x30FC;&#x306E;&#x91CD;&#x8981;&#x306A;&#x52D5;&#x7269;&#x30D4;&#x30B3;&#x30EB;&#x30CA; &#x30A6;&#x30A4;&#x30EB;&#x30B9;&#x53E3;&#x8E44;&#x75AB;&#x30A6;&#x30A4;&#x30EB;&#x30B9; (FMDV) &#x6587;&#x66F8;&#x5316;&#x3055;&#x308C;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x9078;&#x629E;&#x7684;&#x5236;&#x7D04;&#x304C;&#x65E2;&#x306B;&#x305D;&#x306E;&#x9032;&#x5316;&#x306E;&#x524D;&#x306E;&#x6BB5;&#x968E;&#x306E;&#x9593;&#x306B;&#x30A6;&#x30A4;&#x30EB;&#x30B9;&#x4EBA;&#x53E3;&#x3067;&#x7D4C;&#x9A13;&#x3055;&#x308C;&#x3066;&#x3044;&#x308B;&#x305F;&#x3073;&#x306B;&#x30E1;&#x30E2;&#x30EA; &#x30B2;&#x30CE;&#x30E0;&#x306E;&#x30A6;&#x30A4;&#x30EB;&#x30B9;&#x306E;&#x6E96;&#x7A2E;&#x306E;&#x5B58;&#x5728;&#x5F7C;&#x3089;&#x306E;&#x9069;&#x5FDC;&#x5FDC;&#x7B54;&#x52A0;&#x901F;&#x3059;&#x308B;&#x53EF;&#x80FD;&#x6027;&#x304C;&#x3042;&#x308A;&#x307E;&#x3059;&#x3002;&#x5F37;&#x5316;&#x3055;&#x308C;&#x305F;&#x5909;&#x7570; RNA &#x30A6;&#x30A4;&#x30EB;&#x30B9;&#x306E;&#x6570;&#x306E;&#x611F;&#x67D3;&#x6027;&#x306E;&#x640D;&#x5931;&#x306B;&#x3064;&#x306A;&#x304C;&#x308B;&#x3053;&#x3068;&#x304C;&#x793A;&#x3055;&#x308C;&#x3066;&#x3044;&#x308B;: &#x6C34;&#x75B1;&#x6027;&#x53E3;&#x5185;&#x708E;&#x30A6;&#x30A4;&#x30EB;&#x30B9;&#x3001;&#x3072;&#x3068;&#x514D;&#x75AB;&#x4E0D;&#x5168;&#x30A6;&#x30A4;&#x30EB;&#x30B9; 1 &#x578B;&#x30DD;&#x30EA;&#x30AA; &#x30A6;&#x30A4;&#x30EB;&#x30B9;&#x3001;FMDV&#x3002;&#x907A;&#x4F1D;&#x60C5;&#x5831;&#x306E;&#x30E1;&#x30F3;&#x30C6;&#x30CA;&#x30F3;&#x30B9;&#x306E;&#x305F;&#x3081;&#x30B3;&#x30D4;&#x30FC; &#x30A8;&#x30E9;&#x30FC;&#x306E;&#x3057;&#x304D;&#x3044;&#x5024;&#x306E;&#x5B58;&#x5728;&#x306E;&#x7406;&#x8AD6;&#x7684;&#x306A;&#x4E88;&#x6E2C;&#x306B;&#x57FA;&#x3065;&#x304F;&#x3001;&#x3053;&#x308C;&#x3089;&#x306E;&#x89B3;&#x5BDF;&#x306F;&#x3001;&#x65B0;&#x3057;&#x3044;&#x6297;&#x30A6;&#x30A4;&#x30EB;&#x30B9;&#x6226;&#x7565;&#x306E;&#x767A;&#x5C55;&#x306B;&#x8CA2;&#x732E;&#x3059;&#x308B;&#x304B;&#x3082;&#x3057;&#x308C;&#x306A;&#x3044;&#x3002;

&#x51FA;&#x73FE;&#x304A;&#x3088;&#x3073; RNA &#x30A6;&#x30A4;&#x30EB;&#x30B9;&#x306E;&#x4E9C;&#x7A2E;&#x306E;&#x9078;&#x629E;: &#x30E1;&#x30E2;&#x30EA;&#x304A;&#x3088;&#x3073;&#x7D76;&#x6EC5;&#x3002;

&#xBC14;&#xC774;&#xB7EC;&#xC2A4; &#xC131; quasispecies&#xC758; &#xB450; &#xAC00;&#xC9C0; &#xAE30;&#xB2A5; &#xAC80;&#xD1A0;: &#xB3CC;&#xC5F0;&#xBCC0;&#xC774; &#xC2A4;&#xD399;&#xD2B8;&#xB7FC; &#xBC0F; &#xD5A5;&#xC0C1; &#xB41C; mutagenesis &#xC778;&#xD574; &#xBC14;&#xC774;&#xB7EC;&#xC2A4; &#xBA78;&#xC885;&#xC758; &#xC18C;&#xC218; &#xCEF4;&#xD3EC;&#xB10C;&#xD2B8;&#xB85C; &#xBA54;&#xBAA8;&#xB9AC; &#xAC8C;&#xB188;&#xC758; &#xC874;&#xC7AC;. &#xBA54;&#xBAA8;&#xB9AC; &#xC911;&#xC694; &#xD55C; &#xB3D9;&#xBB3C; picornavirus &#xBC1C;-&#xC564;-&#xC785; &#xBCD1; &#xBC14;&#xC774;&#xB7EC;&#xC2A4; (FMDV)&#xC758; &#xC5EC;&#xB7EC; &#xAC00;&#xC9C0; &#xC720;&#xC804;&#xC790; &#xD45C;&#xC2DC;&#xC640; &#xBB38;&#xC11C;&#xD654; &#xB418;&#xC5C8;&#xC2B5;&#xB2C8;&#xB2E4;. &#xBC14;&#xC774;&#xB7EC;&#xC2A4; &#xC131; quasispecies&#xC5D0; &#xBA54;&#xBAA8;&#xB9AC; &#xAC8C;&#xB188;&#xC758; &#xC874;&#xC7AC; &#xB54C;&#xB9C8;&#xB2E4; &#xC120;&#xD0DD;&#xC801; &#xC81C;&#xC57D; &#xC870;&#xAC74; &#xADF8; &#xC9C4;&#xD654;&#xC758; &#xC774;&#xC804; &#xB2E8;&#xACC4; &#xBC14;&#xC774;&#xB7EC;&#xC2A4; &#xC131; &#xC778;&#xAD6C;&#xC5D0; &#xC758;&#xD574; &#xC774;&#xBBF8; &#xACBD;&#xD5D8; &#xD588;&#xB2E4; &#xADF8;&#xB4E4;&#xC758; &#xC801;&#xC751; &#xC751;&#xB2F5;&#xC744; &#xAC00;&#xC18D;&#xD654;&#xD560; &#xC218; &#xC788;&#xC2B5;&#xB2C8;&#xB2E4;. &#xD5A5;&#xC0C1; &#xB41C; mutagenesis infectivity RNA &#xBC14;&#xC774;&#xB7EC;&#xC2A4;&#xC758; &#xC218;&#xC758; &#xC190;&#xC2E4;&#xC744; &#xCD08;&#xB798;&#xD560; &#xD45C;&#xC2DC; &#xB418;&#xC5C8;&#xC2B5;&#xB2C8;&#xB2E4;: &#xC18C;&#xC544;&#xB9C8;&#xBE44;, vesicular &#xAD6C; &#xC5FC; &#xBC14;&#xC774;&#xB7EC;&#xC2A4;, &#xC778;&#xAC04; &#xBA74;&#xC5ED; &#xACB0;&#xD54D; &#xBC14;&#xC774;&#xB7EC;&#xC2A4; 1 &#xD615; &#xBC0F; FMDV. &#xC720;&#xC804; &#xC815;&#xBCF4;&#xC758; &#xC720;&#xC9C0; &#xBCF4;&#xC218;&#xC5D0; &#xB300; &#xD55C; &#xBCF5;&#xC0AC; &#xC624;&#xB958; &#xC784;&#xACC4;&#xAC12;&#xC758; &#xC874;&#xC7AC;&#xC758; &#xC774;&#xB860;&#xC801; &#xC608;&#xCE21;&#xC5D0; &#xB530;&#xB77C; &#xC774;&#xB7EC;&#xD55C; &#xAD00;&#xCE21; &#xD56D; &#xBC14;&#xC774;&#xB7EC;&#xC2A4;&#xC758; &#xC0C8;&#xB85C;&#xC6B4; &#xC804;&#xB7B5;&#xC758; &#xAC1C;&#xBC1C;&#xC5D0; &#xAE30;&#xC5EC;&#xD560; &#xC218; &#xC788;&#xC2B5;&#xB2C8;&#xB2E4;.

&#xCD9C;&#xD604; &#xBC0F; RNA &#xBC14;&#xC774;&#xB7EC;&#xC2A4;&#xC758; &#xBCC0;&#xC885; &#xC911; &#xD558;&#xB098;&#xB97C; &#xC120;&#xD0DD;: &#xBA54;&#xBAA8;&#xB9AC;&#xC640; &#xBA78;&#xC885; &#xC704;&#xAE30;.

Dos caracter&iacute;sticas de quasiespecies virales se repasan: la presencia de los genomas de memoria como componentes minoritarios de sus espectros mutantes y extinci&oacute;n viral por mutag&eacute;nesis mejorada. Memoria se ha documentado con varios marcadores gen&eacute;ticos del virus picornavirus animal importante pie - y - boca disease (VFA). La presencia de los genomas de memoria en quasiespecies virales puede acelerar su respuesta adaptativa siempre que una restricci&oacute;n selectiva ya ha sido experimentada por una poblaci&oacute;n viral durante las etapas anteriores de su evoluci&oacute;n. Mutag&eacute;nesis mejorada se ha demostrado que conducen a la p&eacute;rdida de infectividad de un n&uacute;mero de virus ARN: poliovirus, virus de la estomatitis vesicular, virus de la inmunodeficiencia humana tipo 1 y VFA. Estas observaciones, basadas en la predicci&oacute;n te&oacute;rica de la existencia de un error de copiado-umbral para el mantenimiento de la informaci&oacute;n gen&eacute;tica, pueden contribuir al desarrollo de una nueva estrategia de antiviral.

Aparici&oacute;n y selecci&oacute;n de variantes de virus de ARN: memoria y extinci&oacute;n.

A mutant poliovirus (PV) encoding a change in its polymerase (3Dpol) at a site remote from the catalytic center (G64S) confers reduced sensitivity to ribavirin and forms a restricted quasispecies, because G64S 3Dpol is a high-fidelity enzyme. A foot-and-mouth disease virus (FMDV) mutant that encodes a change in the polymerase catalytic site (M296I) exhibits reduced sensitivity to ribavirin without restricting the viral quasispecies. In order to resolve this apparent paradox, we have established a minimal kinetic mechanism for nucleotide addition by wild-type (WT) FMDV 3Dpol that permits a direct comparison to PV 3Dpol as well as to FMDV 3Dpol derivatives. Rate constants for correct nucleotide addition were on par with those of PV 3Dpol, but apparent binding constants for correct nucleotides were higher than those observed for PV 3Dpol. The A-to-G transition frequency was calculated to be 1/20,000, which is quite similar to that calculated for PV 3Dpol. The analysis of FMDV M296I 3Dpol revealed a decrease in the calculated ribavirin incorporation frequency (1/8,000) relative to that (1/4,000) observed for the WT enzyme. Unexpectedly, the A-to-G transition frequency was higher (1/8,000) than that observed for the WT enzyme. Therefore, FMDV selected a polymerase that increases the frequency of the misincorporation of natural nucleotides while specifically decreasing the frequency of the incorporation of ribavirin nucleotide. These studies provide a mechanistic framework for understanding FMDV 3Dpol structure-function relationships, provide the first direct analysis of the fidelity of FMDV 3Dpol in vitro, identify the beta9-alpha11 loop as a (in)fidelity determinant, and demonstrate that not all ribavirin-resistant mutants will encode high-fidelity polymerases.

Determinants of RNA-dependent RNA polymerase (in)fidelity revealed by kinetic analysis of the polymerase encoded by a foot-and-mouth disease virus mutant with reduced sensitivity to ribavirin.

Armando Arias

Section of Virology

Imperial College London

Reverse Genetics Mediated Recovery of Infectious Murine Norovirus

Armando Arias

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

Imperial College London

Reverse Genetics Mediated Recovery of Infectious Murine Norovirus

Section of Virology