Synthesis of a novel hepatitis C virus protein by ribosomal frameshift

Abstract

Hepatitis C virus (HCV) is an important human pathogen that affects ∼100 million people worldwide. Its RNA genome codes for a polyprotein, which is cleaved by viral and cellular proteases to produce at least 10 mature viral protein products. We report here the discovery of a novel HCV protein synthesized by ribosomal frameshift. This protein, which we named the F protein, is synthesized from the initiation codon of the polyprotein sequence followed by ribosomal frameshift into the −2/+1 reading frame. This ribosomal frameshift requires only codons 8–14 of the core protein‐coding sequence, and the shift junction is located at or near codon 11. An F protein analog synthesized in vitro reacted with the sera of HCV patients but not with the sera of hepatitis B patients, indicating the expression of the F protein during natural HCV infection. This unexpected finding may open new avenues for the development of anti‐HCV drugs.

The HCV ARFP/F/core+1 protein: Production and functional analysis of an unconventional viral product

“…Extensive site-directed mutagenesis within the core/core11 coding region (18) and radioactive sequencing of the protein synthesized in vitro (19) showed that the production of this novel 16 kDa protein resulted from the initiation of translation at the AUG initiator codon of the polyprotein sequence, followed by a 22/11 ribosomal frameshift in an adenosine (A)-rich region located within core codons 8-11. The p16 protein is known as ARFP (alternative reading frame protein) (17), F (for frameshift) (19) or core11 (to indicate the position of the new open reading frame) protein (18,20).…”

“…The p16 protein is known as ARFP (alternative reading frame protein) (17), F (for frameshift) (19) or core11 (to indicate the position of the new open reading frame) protein (18,20).…”

“…Since its discovery, several studies have shown that the core11/ARFP/F protein is specifically recognized by antibodies in serum (17)(18)(19)(33)(34)(35)(36)(37)(38)(39)(40) and by the memory T cells (34,36) of HCV-infected patients, providing evidence that the core11 ORF is produced during natural HCV infection in humans. However, initial studies gave variable frequencies for the detection of core11-humoral immune responses in HCV-infected individuals.…”

“…We subsequently showed, with lysates of E. coli cells producing a recombinant GST-core11 protein from an HCV-1 isolate lacking the first 10 core amino acids [designated D(1-10)core11/F], the detection on western blots of specific antibodies in 77% of the HCV-positive sera from chronically infected patients tested (26 of 34 sera) (18). Xu et al (19) showed that radiolabeled HCV-1 core11/F and D(1-3)core11/ F proteins synthesized in vitro (in RRL) were immunoprecipitated with six and five, respectively, of six HCV-positive serum samples tested, whereas a D(1-3)core11/F antigen expressed in E. coli was recognized by five of eight HCV-positive serum samples in ELISA. The observed variability probably results from differences in the nature of the core11 antigens used in these studies (Fig.…”

“…The existence of an alternative ORF overlapping the core protein gene is inconsistent with the conventional Flaviviridae genetic map, but the HCV core11 ORF is not the only exception. Xu et al (19) have reported the presence of an open reading frame, 109 codons in length, overlapping the GBV-B core gene. The production of a nonstructural protein (Npro) from the amino-terminal structural region has also been reported for the pestivirus genome.…”