1. An analysis of two open reading frames (ORF3 and ORF4) of rat hepatitis E virus genome using its infectious cDNA clones with mutations in ORF3 or ORF4.
- Author
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Tanggis, null, Kobayashi, Tominari, Takahashi, Masaharu, Jirintai, Suljid, Nishizawa, Tsutomu, Nagashima, Shigeo, Nishiyama, Takashi, Kunita, Satoshi, Hayama, Emiko, Tanaka, Takeshi, Mulyanto, null, and Okamoto, Hiroaki
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HEPATITIS E virus , *OPEN reading frames (Genetics) , *VIRAL genomes , *ANTISENSE DNA , *VIRAL replication , *VIRUSES - Abstract
Rat hepatitis E virus (ratHEV) genome has four open reading frames (ORFs: ORF1, ORF2, ORF3 and ORF4). The functions of ORF3 and ORF4 are unknown. An infectious cDNA clone (pUC-ratELOMB-131L_wt, wt) and its derivatives including ORF3-defective (ΔORF3) and ORF4-defective (ΔORF4) mutants, were constructed and their full-length RNA transcripts transfected into PLC/PRF/5 cells. ΔORF3 replicated as efficiently as wt in cells. However, ≤1/1000 of the number of progenies were detectable in the culture supernatant of ΔORF3-infected cells compared with wt-infected cells. ORF4 protein was not detectable in ratHEV-infected cells or in the liver tissues of ratHEV-infected rats. No marked differences were noted between wt and ΔORF4 regarding the viral replication and protein expression. ORF3 mutants with proline-to-leucine mutations at amino acids (aa) 93, 96 and/or 98 in ORF3 were constructed and transfected into PLC/PRF/5 cells. Wt and an ORF3 mutant with leucine at aa 98 (ORF3-L98) replicated efficiently (density 1.15–1.16 g/cm 3 ), while ORF3-L93 + L96 exhibited a decreased viral release and banded at 1.26–1.27 g/cm 3 , similar to ΔORF3. In conclusion, the ORF3 protein, especially its proline residues at aa 93 and 96, is essential for the release of membrane-associated ratHEV particles, and ORF4 is unnecessary for the replication of ratHEV. [ABSTRACT FROM AUTHOR]
- Published
- 2018
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