Endoplasmic Reticulum Stress Induced Synthesis of a Novel Viral Factor Mediates Efficient Replication of Genotype-1 Hepatitis E Virus

Abstract

Hepatitis E virus (HEV) causes acute hepatitis in many parts of the world including Asia, Africa and Latin America. Though self-limiting in normal individuals, it results in ~30% mortality in infected pregnant women. It has also been reported to cause acute and chronic hepatitis in organ transplant patients. Of the seven viral genotypes, genotype-1 virus infects humans and is a major public health concern in South Asian countries. Sporadic cases of genotype-3 and 4 infection in human and animals such as pigs, deer, mongeese have been reported primarily from industrialized countries. Genotype-5, 6 and 7 viruses are known to infect animals such as wild boar and camel, respectively. Genotype-3 and 4 viruses have been successfully propagated in the laboratory in mammalian cell culture. However, genotype-1 virus replicates poorly in mammalian cell culture and no other efficient model exists to study its life cycle. Here, we report that endoplasmic reticulum (ER) stress promotes genotype-1 HEV replication by inducing cap-independent, internal initiation mediated translation of a novel viral protein (named ORF4). Importantly, ORF4 expression and stimulatory effect of ER stress inducers on viral replication is specific to genotype-1. ORF4 protein sequence is mostly conserved among genotype-1 HEV isolates and ORF4 specific antibodies were detected in genotype-1 HEV patient serum. ORF4 interacted with multiple viral and host proteins and assembled a protein complex consisting of viral helicase, RNA dependent RNA polymerase (RdRp), X, host eEF1α1 (eukaryotic elongation factor 1 isoform-1) and tubulinβ. In association with eEF1α1, ORF4 stimulated viral RdRp activity. Furthermore, human hepatoma cells that stably express ORF4 or engineered proteasome resistant ORF4 mutant genome permitted enhanced viral replication. These findings reveal a positive role of ER stress in promoting genotype-1 HEV replication and pave the way towards development of an efficient model of the virus.

Hepatitis E virus replication and interferon responses in human placental cells

“…The infection with primary material of the Sar55 genotype 1 isolate was restricted to human liver cells, but the infection efficiency was limited. Recently, Nair et al51 described a novel protein, termed ORF4, which is gt1 specific and implicated in the endoplasmic stress response promoting viral replication. If ORF4 is expressed during pregnancy in HEV infection in vivo and how this protein interacts with the host response require future investigation.…”

Review article: hepatitis E—a concise review of virology, epidemiology, clinical presentation and therapy

“…11 However, ORF4 is unique domain of unknown function, a helicase (Hel) domain, and a RNAdependent RNA polymerase (RdRp). 18 However, the protease activity of the PCP domain is a subject of some debate, with some reports suggesting that this domain may instead de-ubiquinate proteins, preventing the prosomal degradation of protein required for viral replication.…”

Detection of HEV in naturally infected swine from central Argentina by an optimized HEV/MS2 duplex RT‐qPCR

“…HEV strains isolated from human, and other animal reservoirs, belong to Orthohepevirus A species, have been classified into eight distinct genotypes (HEV‐1 to HEV‐8) (Smith et al, ; Sridhar, Teng, Chiu, Lau, & Woo, ) and further divided into several subtypes (Lu, Li, & Hagedorn, ; Smith et al, ). Most genotypes encode three overlapping open reading frames (ORF) ORF1, ORF2 and ORF3, whereas an additional ORF4 was reported for HEV‐1 (Emerson & Purcell, ; Lu et al, ; Nair et al, ). HEV‐3 is the most prevalent in industrialized countries of Europe and America being proposed as zoonotically transmitted to humans or via the environment.…”