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Host heterogeneous ribonucleoprotein K (hnRNP K) as a potential target to suppress hepatitis B virus replication.

ABSTRACT: BACKGROUND: Hepatitis B virus (HBV) infection results in complications such as cirrhosis and hepatocellular carcinoma. Suppressing viral replication in chronic HBV carriers is an effective approach to controlling disease progression. Although antiviral compounds are available, we aimed to identify host factors that have a significant effect on viral replication efficiency. METHODS AND FINDINGS: We studied a group of hepatitis B carriers by associating serum viral load with their respective HBV genomes, and observed a significant association between high patient serum viral load with a natural sequence variant within the HBV enhancer II (Enh II) regulatory region at position 1752. Using a viral fragment as an affinity binding probe, we isolated a host DNA-binding protein belonging to the class of heterogeneous nuclear ribonucleoproteins--hnRNP K--that binds to and modulates the replicative efficiency of HBV. In cell transfection studies, overexpression of hnRNP K augmented HBV replication, while gene silencing of endogenous hnRNP K carried out by small interfering RNAs resulted in a significant reduction of HBV viral load. CONCLUSION: The evidence presented in this study describes a wider role for hnRNP K beyond maintenance of host cellular functions and may represent a novel target for pharmacologic intervention of HBV replication.

SUBMITTER: Ng LF

PROVIDER: S-EPMC1181871 | biostudies-literature | 2005 Jul

REPOSITORIES: biostudies-literature

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Host heterogeneous ribonucleoprotein K (hnRNP K) as a potential target to suppress hepatitis B virus replication.

Ng Lisa F P LF Chan Marieta M Chan Soh-Ha SH Cheng Paul Chung-Pui PC Leung Eastwood Hon-Chiu EH Chen Wei-Ning WN Ren Ee-Chee EC

PLoS medicine 20050726 7

<h4>Background</h4>Hepatitis B virus (HBV) infection results in complications such as cirrhosis and hepatocellular carcinoma. Suppressing viral replication in chronic HBV carriers is an effective approach to controlling disease progression. Although antiviral compounds are available, we aimed to identify host factors that have a significant effect on viral replication efficiency.<h4>Methods and findings</h4>We studied a group of hepatitis B carriers by associating serum viral load with their res ...[more]

PMID: 16033304

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Project description:Serum from a patient showing symptoms related to autoimmunity was found to contain autoantibodies to the nuclear mitotic apparatus (NuMA) protein and to several novel nuclear antigens with estimated molecular weights of 40, 43, 72, 74 and 82 kDa. Using this serum for screening a human cDNA expression library a 2.5 kb cDNA clone was isolated which encoded the complete sequence of a protein of 633 amino acids. Sequence analysis revealed a modular structure of the protein: an acidic N-terminal region of approximately 150 amino acids was followed by three adjacent consensus sequence RNA binding domains located in the central part of the protein. In the C-terminal portion a nuclear localization signal and an octapeptide (PPPRMPPP) with similarity to a major B cell epitope of the snRNP core protein B were identified. This was followed by a glycine- and arginine-rich section of approximately 120 amino acids forming another type of RNA binding motif, a RGG box. Interestingly, three copies of a tyrosine-rich decapeptide were found interspersed in the RGG box region. The major in vitro translation product of the cDNA co-migrated in SDS-PAGE with the 82 kDa polypeptide that was recognized by autoantibodies. The structural motifs as well as the immunofluorescence pattern generated by anti-82 kDa antibodies suggested that the antigen was one of the proteins of the heterogeneous nuclear ribonucleoprotein (hnRNP) complex. Subsequently the 82 kDa antigen was identified as hnRNP R protein by its presence in immunoprecipitated hnRNP complexes and co-migration of the recombinant protein with this hitherto uncharacterized hnRNP constituent in two-dimensional gel electrophoresis. The concomitant autoimmune response to a hnRNP component of the pre-mRNA processing machinery and to NuMA, a protein engaged in mitotic events and reported to be associated with mRNA splicing complexes in interphase, may indicate physical and functional association of these antigens. Support for this notion comes from observations that concomitant or coupling of autoantibody responses to proteins which are associated with each other as components of subcellular particles are often found in autoimmune diseases.

Project description:The mechanism underlying the sensing of varying degrees of physiological folate deficiency, prior to adaptive optimization of cellular folate uptake through the translational up-regulation of folate receptors (FR) is unclear. Because homocysteine, which accumulates intracellularly during folate deficiency, stimulated interactions between heterogeneous nuclear ribonucleoprotein E1 (hnRNP-E1) and an 18-base FR-? mRNA cis-element that led to increased FR biosynthesis and net up-regulation of FR at cell surfaces, hnRNP-E1 was a plausible candidate sensor of folate deficiency. Accordingly, using purified components, we evaluated the physiological basis whereby L-homocysteine triggered these RNA-protein interactions to stimulate FR biosynthesis. L-homocysteine induced a concentration-dependent increase in RNA-protein binding affinity throughout the range of physiological folate deficiency, which correlated with a proportionate increase in translation of FR in vitro and in cultured human cells. Targeted reduction of newly synthesized hnRNP-E1 proteins by siRNA to hnRNP-E1 mRNA reduced both constitutive and L-homocysteine-induced rates of FR biosynthesis. Furthermore, L-homocysteine covalently bound hnRNP-E1 via multiple protein-cysteine-S-S-homocysteine mixed disulfide bonds within K-homology domains known to interact with mRNA. These data suggest that a concentration-dependent, sequential disruption of critical cysteine-S-S-cysteine bonds by covalently bound L-homocysteine progressively unmasks an underlying RNA-binding pocket in hnRNP-E1 to optimize interaction with FR-? mRNA cis-element preparatory to FR up-regulation. Collectively, such data incriminate hnRNP-E1 as a physiologically relevant, sensitive, cellular sensor of folate deficiency. Because diverse mammalian and viral mRNAs also interact with this RNA-binding domain with functional consequences to their protein expression, homocysteinylated hnRNP-E1 also appears well positioned to orchestrate a novel, nutrition-sensitive (homocysteine-responsive), posttranscriptional RNA operon in folate-deficient cells.

Dataset Information

Host heterogeneous ribonucleoprotein K (hnRNP K) as a potential target to suppress hepatitis B virus replication.

Publications

Host heterogeneous ribonucleoprotein K (hnRNP K) as a potential target to suppress hepatitis B virus replication.

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