2 research outputs found
Role of the cellular decapping activator LSM1-7 complex in the replication of positive-strand RNA viruses
By using the ability of the positive-strand RNA ((+)RNA) virus BMV to replicate in yeast it was previously shown that subunits of the LSm1-7 ring, as well as Pat1 and Dhh1 play an essential role in the transit of the BMV genome from translation to replication. In non-infected cells, these proteins mediate the transition of cellular mRNAs from a translational to a non-translational state by activating decapping in the 5'-3' - deadenylation-dependent mRNA decay pathway. Given the conservation of this pathway from yeast to humans and the common need of all (+)RNA viruses to regulate the transition of their genomes from active translation to a translationally inactive state to allow replication, an exciting possibility, and our working hypothesis, was that LSm1-7, Dhh1 and Pat1 are used not only by BMV to replicate in yeast but also by human (+) RNA viruses, such as HCV, to replicate in mammalian cells. Furthermore, given the key role of these proteins in a common step to all (+)RNA viruses, it is essential to characterize the not yet defined molecular mechanisms associated with such function. In this regard, we also hypothesized that the LSm1-7 complex, as member of the Sm family of proteins, would directly interact with viral genomes of (+)RNA viruses in order to play their role in the virus life cycle in a similar way that other family counterparts directly interact with their RNA targets in order to achieve their different cellular functions. In this work we were able to confirm both hypothesis showing that human homologues of the upper mentioned proteins LSm1-7, Rck/p54 and PatL1, are required for HCV RNA translation and replication. Additionally, we also showed that reconstituted LSm1-7 complexes specifically recognize important signals, either in BMV or HCV genomes, that regulate their translation and/or replication. These observations constitute the first evidence that the LSm1-7 complex is able to directly interact with viral genomes representing also novel LSm1-7 interaction sites. Given the common replication strategies of (+)RNA viruses and the conserved cellular functions of LSm1-7, Pat1 and Dhh1 from yeast to humans, our findings pinpoint a weak spot that may be exploited to generate broad-spectrum antiviral drugs.Utilizando la capacidad del BMV, un virus de ARN de cadena positiva (ARN(+)), para replicar en levaduras se ha demostrado previamente que las subunidades del anillo LSm1-7, as铆 como Pat1 y Dhh1, desempe帽an un papel esencial en la transici贸n del genoma del virus de BMV desde traducci贸n a replicaci贸n. En c茅lulas no infectadas, estas prote铆nas median la transici贸n de ARNm celulares de la traducci贸n a un estado de no-traducci贸n mediante la activaci贸n del proceso de decapping en la via 5'-3' de degradaci贸n de los ARNs celulares dependiente de deadenilaci贸n. Teniendo en cuenta la conservaci贸n de esta v铆a desde levaduras a humanos y la necesidad com煤n de todos los virus ARN(+) para regular la transici贸n de sus genomas desde un estado activo de traducci贸n a otro no activo para permitir la replicaci贸n, una posibilidad interesante, y nuestra hip贸tesis de trabajo, es que LSm1-7, Dhh1 y Pat1 son utilizadas no solo por BMV para replicar en levaduras, sino tambi茅n por otros virus ARN(+) que infectan a humanos, como el virus de la hepatitis C, para replicar en c茅lulas de mam铆feros. Por otra parte, dado el papel clave de estas prote铆nas en un paso com煤n en todos los virus de ARN(+), es esencial caracterizar los mecanismos moleculares aun no conocidos y asociados a dicha funci贸n. En este sentido, tambi茅n estudiamos la hip贸tesis de que el complejo LSm1-7, como miembro de la familia de prote铆nas Sm, pueda interactuar directamente con los genomas virales de virus de ARN(+) con el fin de desempe帽ar su papel en el ciclo de vida del virus de una manera similar a la que otros miembros de su familia interact煤an con sus ARN con el fin de lograr sus diferentes funciones celulares. En este trabajo hemos podido confirmar ambas hip贸tesis demostrando que los hom贸logos humanos de las prote铆nas anteriormente mencionadas, LSm1-7, Rck/p54 y PatL1, son necesarios para la traducci贸n y replicaci贸n del ARN del virus de la Hepatitis C. Por otra parte, los anillos reconstituidos de LSm1-7 reconocen espec铆ficamente se帽ales importantes, tanto en el genoma de BMV como en el de la Hepatitis C que regulan su traducci贸n y/o replicaci贸n. Estas observaciones constituyen la primera evidencia de que el complejo LSm1-7 es capaz de interactuar directamente con genomas virales y representan tambi茅n novedosos patrones de interacci贸n de este complejo con ARN. Teniendo en cuenta las estrategias de replicaci贸n en com煤n de los virus de ARN de cadena positiva y las funciones celulares conservadas de LSm1-7, Pat1 y Dhh1 de levaduras a humanos, nuestros resultados se帽alan la posibilidad de explotar estas prote铆nas para la generaci贸n de medicamentos antivirales de amplio espectro
LSm1-7 complexes bind to specific sites in viral RNA genomes and regulate their translation and replication
LSm1-7 complexes promote cellular mRNA degradation, in addition to translation and replication of positive-strand RNA viruses such as the Brome mosaic virus (BMV). Yet, how LSm1-7 complexes act on their targets remains elusive. Here, we report that reconstituted recombinant LSm1-7 complexes directly bind to two distinct RNA-target sequences in the BMV genome, a tRNA-like structure at the 3'-untranslated region and two internal A-rich single-stranded regions. Importantly, in vivo analysis shows that these sequences regulate the translation and replication of the BMV genome. Furthermore, both RNA-target sequences resemble those found for Hfq, the LSm counterpart in bacteria, suggesting conservation through evolution. Our results provide the first evidence that LSm1-7 complexes interact directly with viral RNA genomes and open new perspectives in the understanding of LSm1-7 functions.This work was supported by grants from the Spanish Ministerio de Educaci贸n y Ciencia (BFU2007-66933/BMC) and the German Research Foundation (DFG-FOR855). I.A.-R. and D.L. were supported by Funda莽ao para a Ci锚ncia e Tecnolog铆a (SARH/BD/9630/2002; SFRH/BD/37047/2007