Dataset Information

Identification and characterization of a hepatitis delta virus RNA transcriptional promoter.

ABSTRACT: Transcription and replication of hepatitis delta virus (HDV) RNA are performed by the cellular enzyme RNA polymerase II (Pol II). As DNA is the normal template for Pol II, the enzyme must undergo template switching. The mechanism for this is unknown, but since HDV RNA can form a rod-like molecule by extensive intramolecular base pairing, it has been suggested that a double-stranded region(s) of HDV RNA serves as a recognition site for Pol II. A bidirectional promoter has been identified previously on HDV cDNA (T. B. Macnaughton, M. R. Beard, M. Chao, E. J. Gowans, and M. M. C. Lai, Virology 196:629-636, 1993). In the present study, genomic RNA corresponding to this region was able to direct the synthesis of antigenomic RNA in a nuclear extract transcription reaction, whereas genomic RNA species containing a mutation that resulted in a replication-defective phenotype were unable to do so. Thus, this region, the location of which is defined as nucleotides 1608 to 1669 on the basis of a highly conserved structure, represents a RNA-RNA promoter. Computer analysis of the RNA secondary structure predicted that the promoter contains two bulge regions in a stem-loop structure which encompasses a GC-rich motif. This predicted model was confirmed by enzyme digestion and primer extension analysis. The promoter is located at one end of the rod and has some homology with the simian virus 40 late promoter. A number of other mutations were introduced within this region, and expression plasmids were constructed to examine the effects of mutations in the promoter on HDV replication. Disruption of the overall secondary structure, particularly the bulge regions, totally inhibited HDV RNA replication.

SUBMITTER: Beard MR

PROVIDER: S-EPMC190452 | biostudies-other | 1996 Aug

REPOSITORIES: biostudies-other

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Identification and characterization of a hepatitis delta virus RNA transcriptional promoter.

Beard M R MR MacNaughton T B TB Gowans E J EJ

Journal of virology 19960801 8

Transcription and replication of hepatitis delta virus (HDV) RNA are performed by the cellular enzyme RNA polymerase II (Pol II). As DNA is the normal template for Pol II, the enzyme must undergo template switching. The mechanism for this is unknown, but since HDV RNA can form a rod-like molecule by extensive intramolecular base pairing, it has been suggested that a double-stranded region(s) of HDV RNA serves as a recognition site for Pol II. A bidirectional promoter has been identified previous ...[more]

PMID: 8764005

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Project description:Hepatitis delta virus (HDV) is a satellite of hepatitis B virus that increases the severity of acute and chronic liver disease. HDV produces three processed RNAs that accumulate in infected cells: the circular genome; the circular antigenome, which serves as a replication intermediate; and lesser amounts of the mRNA, which encodes the sole viral protein, hepatitis delta antigen (HDAg). The HDV genome and antigenome RNAs form ribonucleoprotein complexes with HDAg. Although HDAg is required for HDV replication, it is not known how the relative amounts of HDAg and HDV RNA affect replication, or whether HDAg synthesis is regulated by the virus. Using a novel transfection system in which HDV replication is initiated using in vitro-synthesized circular HDV RNAs, HDV replication was found to depend strongly on the relative amounts of HDV RNA and HDAg. HDV controls these relative amounts via differential effects of HDAg on the production of HDV mRNA and antigenome RNA, both of which are synthesized from the genome RNA template. mRNA synthesis is favored at low HDAg levels but becomes saturated at high HDAg concentrations. Antigenome RNA accumulation increases linearly with HDAg and dominates at high HDAg levels. These results provide a conceptual model for how HDV antigenome RNA production and mRNA transcription are controlled from the earliest stage of infection onward and also demonstrate that, in this control, HDV behaves similarly to other negative-strand RNA viruses, even though there is no genetic similarity between them.IMPORTANCE Hepatitis delta virus (HDV) is a satellite of hepatitis B virus that increases the severity of liver disease; approximately 15 million people are chronically infected worldwide. There are no licensed therapies available. HDV is not related to any known virus, and few details regarding its replication cycle are known. One key question is whether and how HDV regulates the relative amounts of viral RNA and protein in infected cells. Such regulation might be important because the HDV RNA and protein form complexes that are essential for HDV replication, and the proper stoichiometry of these complexes could be critical for their function. Our results show that the relative amounts of HDV RNA and protein in cells are indeed important for HDV replication and that the virus does control them. These observations indicate that further study of these regulatory mechanisms is required to better understand replication of this serious human pathogen.

Dataset Information

Identification and characterization of a hepatitis delta virus RNA transcriptional promoter.

Publications

Identification and characterization of a hepatitis delta virus RNA transcriptional promoter.

Shared Molecules

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0.5

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