Identification of AP80978, a novel small-molecule inhibitor of hepatitis C virus replication that targets NS4B.

A small-molecule inhibitor of hepatitis C virus (HCV) designated AP89652 was identified by screening a compound library with an HCV genotype 1b subgenomic replicon assay. AP89652 contains two chiral centers, and testing of two syn enantiomers revealed that activity in the replicon assay resided with only one, AP80978, whose 50% effective concentration (EC50) (the concentration at which a 50% reduction in Renilla luciferase levels was observed relative to an untreated control) was 630 nM. AP80978 was inhibitory against HCV genotypes 1a and 1b but not genotype 2a.

Translation and replication of hepatitis C virus genomic RNA depends on ancient cellular proteins that control mRNA fates.

Inevitably, viruses depend on host factors for their multiplication. Here, we show that hepatitis C virus (HCV) RNA translation and replication depends on Rck/p54, LSm1, and PatL1, which regulate the fate of cellular mRNAs from translation to degradation in the 5'-3'-deadenylation-dependent mRNA decay pathway. The requirement of these proteins for efficient HCV RNA translation was linked to the 5' and 3' untranslated regions (UTRs) of the viral genome.

The DEAD-box RNA helicase Ded1p affects and accumulates in Saccharomyces cerevisiae P-bodies.

Recent results suggest that cytoplasmic mRNAs can form translationally repressed messenger ribonucleoprotein particles (mRNPs) capable of decapping and degradation, or accumulation into cytoplasmic processing bodies (P-bodies), which can function as sites of mRNA storage. The proteins that function in transitions between the translationally repressed mRNPs that accumulate in P-bodies and mRNPs engaged in translation are largely unknown. Herein, we demonstrate that the yeast translation initiation factor Ded1p can localize to P-bodies.

Identification of novel small-molecule inhibitors of West Nile virus infection.

West Nile virus (WNV) has spread throughout the United States and Canada and now annually causes a clinical spectrum of human disease ranging from a self-limiting acute febrile illness to acute flaccid paralysis and lethal encephalitis. No therapy or vaccine is currently approved for use in humans. Using high-throughput screening assays that included a luciferase expressing WNV subgenomic replicon and an NS1 capture enzyme-linked immunosorbent assay, we evaluated a chemical library of over 80,000 compounds for their capacity to inhibit WNV replication.

Host deadenylation-dependent mRNA decapping factors are required for a key step in brome mosaic virus RNA replication.

The genomes of positive-strand RNA [+RNA] viruses perform two mutually exclusive functions: they act as mRNAs for the translation of viral proteins and as templates for viral replication. A universal key step in the replication of +RNA viruses is the coordinated transition of the RNA genome from the cellular translation machinery to the viral replication complex. While host factors are involved in this step, their nature is largely unknown.

Detecting differential gene expression with a semiparametric hierarchical mixture method.

Mixture modeling provides an effective approach to the differential expression problem in microarray data analysis. Methods based on fully parametric mixture models are available, but lack of fit in some examples indicates that more flexible models may be beneficial. Existing, more flexible, mixture models work at the level of one-dimensional gene-specific summary statistics, and so when there are relatively few measurements per gene these methods may not provide sensitive detectors of differential expression.

Systematic, genome-wide identification of host genes affecting replication of a positive-strand RNA virus.

Positive-strand RNA viruses are the largest virus class and include many pathogens such as hepatitis C virus and the severe acute respiratory syndrome coronavirus (SARS). Brome mosaic virus (BMV) is a representative positive-strand RNA virus whose RNA replication, gene expression, and encapsidation have been reproduced in the yeast Saccharomyces cerevisiae. By using traditional yeast genetics, host genes have been identified that function in controlling BMV translation, selecting BMV RNAs as replication templates, activating the replication complex, maintaining a lipid composition required for membrane-associated RNA replication, and other steps.

Yeast Lsm1p-7p/Pat1p deadenylation-dependent mRNA-decapping factors are required for brome mosaic virus genomic RNA translation.

Previously, we used the ability of the higher eukaryotic positive-strand RNA virus brome mosaic virus (BMV) to replicate in yeast to show that the yeast LSM1 gene is required for recruiting BMV RNA from translation to replication. Here we extend this observation to show that Lsm1p and other components of the Lsm1p-Lsm7p/Pat1p deadenylation-dependent mRNA decapping complex were also required for translating BMV RNAs. Inhibition of BMV RNA translation was selective, with no effect on general cellular translation.

Brome mosaic virus RNA replication: revealing the role of the host in RNA virus replication.

The replication of positive-strand RNA viruses is a complex multi-step process involving interactions between the viral genome, virus-encoded replication factors, and host factors. The plant virus brome mosaic virus (BMV) has served as a model for positive-strand RNA virus replication, recombination, and virion assembly. This review addresses recent findings on the identification and characterization of host factors in BMV RNA replication.

An alternate pathway for recruiting template RNA to the brome mosaic virus RNA replication complex.

The multidomain RNA replication protein 1a of brome mosaic virus (BMV), a positive-strand RNA virus in the alphavirus-like superfamily, plays key roles in assembly and function of the viral RNA replication complex. 1a, which encodes RNA capping and helicase-like domains, localizes to endoplasmic reticulum membranes, recruits BMV 2a polymerase and viral RNA templates, and forms membrane-bound, capsid-like spherules in which RNA replication occurs. cis-acting signals necessary and sufficient for RNA recruitment by 1a have been mapped in BMV genomic RNA2 and RNA3.