Enterovirus 71 (EV71) is a major causative agent of hand, foot and mouth disease (HFMD), a common febrile disease occurring mainly in young children. Although clinical manifestations of HFMD are usually mild and self limiting, a severe EV71 outbreak can lead to a diverse array of neurological diseases. Identification of the specific cellular receptors is crucial for elucidating the mechanism of early virus-host interactions and the pathogenesis of enteroviruses. Here we identify human P-selectin glycoprotein ligand-1 (PSGL-1; CD162), a sialomucin membrane protein expressed on leukocytes that has a major role in early stages of inflammation, as a functional receptor for EV71 using an expression cloning method by panning. The N-terminal region of PSGL-1 binds specifically to EV71. Stable PSGL-1 expression allowed EV71 entry and replication, and development of cytopathic effects in nonsusceptible mouse L929 cells. Five out of eight EV71 strains bound soluble PSGL-1 and used intact PSGL-1 as the primary receptor for infection of Jurkat T cells. Three other EV71 strains did not use PSGL-1, suggesting the presence of strain-specific replication of EV71 in leukocytes. EV71 replicated in nonleukocyte cell lines in a PSGL-1-independent manner, indicating the presence of alternative receptor(s) for EV71. The identification of PSGL-1 as a receptor for EV71 sheds new light on a role for PSGL-1-positive leukocytes in cell tropism and pathogenesis during the course of HFMD and other EV71-mediated diseases.

No abstract

The hepatitis C virus (HCV) nonstructural protein (NS)5AHepatitis C virus (HCV) infects 170 million people worldwide and frequently leads to cirrhosis or hepatocellular carcinoma (6, 29). HCV is classified in the family Flaviviridae and possesses a single-stranded positive-sense RNA with a length of 9.6 kb. The HCV genome encodes a single large precursor polyprotein composed of about 3,000 amino acids (aa) that is processed by cellular and viral proteases, resulting in at least 10 structural and nonstructural (NS) proteins (29). Details of HCV's replication cycle are unknown because of the low viral load in the sera of HCV-infected individuals and the lack of a reliable and robust cell culture system to support HCV infection and replication. The development of HCV RNA replicons in which a synthetic HCV genomic or subgenomic RNA replicates efficiently in the human hepatocarcinoma cell line Huh-7 has enabled the study of viral RNA replication in cell culture (4,20,24). The HCV RNA replication complex, composed of the viral NS proteins and host cellular proteins, replicates the viral RNA genome at the intracellular membrane. Thus far, the HCV replicon system has greatly contributed to the understanding of HCV replication and pathogenesis associated with the expression of viral NS proteins. Replication of positive-strand RNA viruses generally involves certain intracellular membrane structures, including the endoplasmic reticulum (ER), Golgi apparatus, endosome, and lysosome (39).Recently, several groups have succeeded in demonstrating cellfree replication activities of replication complexes in crude membrane fractions of HCV subgenomic replicon cells (2, 3, 14, 53). These cell-free systems provide semi-intact polymerase assays for biochemical dissection of HCV RNA replication and are a useful source for the isolation of HCV replication complexes. Replication complexes were detected in detergent-resistant membrane structures, most likely lipid raft structures (2, 14). Although HCV NS proteins presumably form a membrane-associated RNA replication complex with host proteins, the precise components and mechanisms for replication are poorly understood.HCV NS5A is a phosphoprotein that appears to possess multiple and diverse functions in viral replication, interferon resistance, and pathogenesis (26,35). Cell culture-adaptive mutations have been shown to cluster in the central portion of NS5A in subgenomic HCV replicons, indicating that NS5A is involved in the viral replication process either directly or by interacting with host cellular proteins (4, 55). This observation, together with the modulation of NS5A hyperphosphorylation by NS3, NS4A, and NS4B and physical interaction with other viral NS proteins, strongly supports the notion that NS5A is an essential component of the HCV replication complex (21,30,36). NS5A has been shown to be associated with a range of cellular proteins involved in cellular signaling pathways, such as interferon-induced kinase PKR (11), growth factor receptor-binding protein 2 (Grb2) (45), p53...

Hepatitis C virus (HCV) nonstructural protein 5A (NS5A) is a component of viral replicase and is well known to modulate the functions of several host proteins. Here, we show that NS5A specifically interacts with FKBP8, a member of the FK506-binding protein family, but not with other homologous immunophilins. Three sets of tetratricopeptide repeats in FKBP8 are responsible for interactions with NS5A. The siRNA-mediated knockdown of FKBP8 in a human hepatoma cell line harboring an HCV RNA replicon suppressed HCV RNA replication, and this reduction was reversed by the expression of an siRNA-resistant FKBP8 mutant. Furthermore, immunoprecipitation analyses revealed that FKBP8 forms a complex with Hsp90 and NS5A. Treatment of HCV replicon cells with geldanamycin, an inhibitor of Hsp90, suppressed RNA replication in a dose-dependent manner. These results suggest that the complex consisting of NS5A, FKBP8, and Hsp90 plays an important role in HCV RNA replication.