The tiny hepatitis delta virus (HDV) antigen (SHDAg) plays an essential

The tiny hepatitis delta virus (HDV) antigen (SHDAg) plays an essential role in HDV RNA double-rolling-circle replication. S177 and mass spectrometric analysis. Interestingly we found an increase in the HDV replication from antigenomic RNA to genomic RNA but not in that from genomic RNA to antigenomic RNA. The Ser-177 residue was critical for SHDAg conversation with RNA polymerase II (RNAPII) the enzyme proposed to regulate antigenomic RNA replication. These results demonstrate the role of ERK1/2-mediated Ser-177 phosphorylation in modulating HDV antigenomic RNA replication possibly through RNAPII regulation. The results may shed light on the mechanisms of HDV RNA replication. Hepatitis delta computer virus (HDV) is usually a subviral pathogen that can self-replicate but relies on its helper hepatitis B computer virus to provide envelope proteins for HDV contamination and virion assembly (3). HDV is usually a negative-stranded RNA computer virus with 1.7-kb single-stranded circular RNA (genomic RNA) that because of the intramolecular base pairing is usually folded into an unbranched rod-like structure (11 53 The genomic strand of HDV RNA does not encode any protein but the complementary strand (antigenomic HDV RNA) which replicates from genomic RNA encodes hepatitis AEG 3482 delta antigen (HDAg) (53). You will find two forms of HDAg: p24 (small HDAg [SHDAg]) and p27 (large HDAg [LHDAg]) (54). SHDAg is usually translated from a 0.8-kb subantigenomic message RNA that transcribes from HDV genomic RNA (19). SHDAg is essential for HDV genomic and antigenomic RNA replication (11 35 During HDV replication the cells accumulate multimeric HDV genomic or antigenomic RNA which AEG 3482 exhibit ribozyme activity for autocatalytic cleavage and self-ligation to form monomeric circular RNAs (1 5 28 44 56 In the late stage of the viral replication cycle the LHDAg is definitely synthesized after antigenomic RNA editing (45 51 LHDAg shares the 195-amino-acid sequence with SHDAg except the C terminus of LHDAg has an additional 19-amino-acid extension (62). AEG 3482 The function of LHDAg is definitely important for packaging with hepatitis B computer virus surface antigen to form the adult HDV computer virus particle (8 15 24 Whether cellular RNA polymerase (RNAP) can result in HDV replication is definitely controversial. John Taylor’s group thought that the HDV replication machinery is definitely carried only by α-amanitin-sensitive RNAPII for viral RNA double-rolling-circle replication (9). However Michael Lai’s group proposed that HDV replication is definitely carried out by two different replication machineries in cells (21 37 39 The first is α-amanitin-sensitive FGF3 RNAP which helps HDV replication from antigenomic RNA to genomic RNA and the additional is definitely α-amanitin-resistant RNAP which processes HDV replication from genomic RNA to antigenomic RNA. The molecular function of SHDAg is essential for HDV double-rolling-circle replication. How SHDAg distinguishes the two phases of viral RNA replication is an interesting query. In addition several posttranslational modifications have been found on HDAgs and these play an essential part in AEG 3482 HDV replication and computer virus packaging (2 15 20 33 40 61 Our earlier studies found that Ser-177 of SHDAg AEG 3482 is definitely a conserved phosphorylation residue that is important for HDV replication from antigenomic RNA to genomic RNA (10 40 41 In addition a mutant SHDAg whose Ser-177 is definitely replaced by alanine cannot be phosphorylated and loses the ability to support the α-amanitin-sensitive HDV replication from antigenomic RNA to the genomic RNA strand (40). Investigating the kinase action on SHDAg at Ser-177 is a good approach to exploring the AEG 3482 mechanism of HDV antigenomic RNA replication. A earlier report recognized RNA-activated protein kinase (PKR) as the kinase responsible for the phosphorylation of SHDAg Ser-177 in vitro and in vivo (10). However PKR appears to be dispensable and fails to upregulate HDV replication in vivo by increasing the phosphorylation of SHDAg at Ser-177. The additional kinase or kinases responsible for SHDAg phosphorylation involved in HDV replication have not been recognized. The aim of our study was to identify the cellular kinase(s) that phosphorylates SHDAg at Ser-177 and regulates HDV replication. The SHDAg Ser-177 is located within the PXS/TP sequence which is a core consensus sequence phosphorylated by proline-directed kinases such as cyclin-dependent kinases (CDKs) and mitogen-activated protein kinases (MAPKs) (43). We looked a bioinformatics database (The Scansite phosphorylation site http://scansite.mit.edu/) which suggested.

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