The 9-kDa hydrophobic protein encoded at the 3′ end of the porcine transmissible gastroenteritis coronavirus genome is membrane-associated

Abstract

The open reading frame potentially encoding a 78 amino acid, 9101 Da hydrophobic protein (HP) and, mapping at the 3' end of the porcine transmissible gastroenteritis coronavirus (TGEV) genome, was shown to be expressed during virus replication. The cloned HP gene was placed in a plasmid under control of the T7 RNA polymerase promoter and in vitro translation of transcripts generated in vitro yielded a 9.1-kDa protein that was immunoprecipitable with porcine hyperimmune anti-TGEV serum. Antiserum raised in rabbits against a 31 amino acid synthetic polypeptide that represented the central hydrophilic region of HP specifically immunoprecipitated HP from TGEV-infected cells. HP was further shown to become associated with microsomal membranes during synthesis in vitro and was found to be closely associated with the endoplasmic reticulum and cell surface membranes in infected cells. The intracellular location of HP suggests that it may play a role in the membrane association of replication complexes or in virion assembly.

Sequence determination of the nucleocapsid protein gene of the porcine epidemic diarrhoea virus confirms that this virus is a coronavirus related to human coronavirus 229E and porcine transmissible gastroenteritis virus

“…The PEDV ORF resembles the BCV I gene in coding for a high leucine content in the predicted products and having the ATG start codon in the same position with respect to that of the N gene. The viruses TGEV, PRCV, FIPV, FECV and CCV possess no such ORF, implying either that the I protein provides a non-essential function or that this function is provided by another protein, for example one encoded by a gene located downstream from the N gene in these viruses, which encodes a hydrophobic product of 78 amino acids (Garwes et al, 1989;Tung et al, 1992).…”

Strategy for Systematic Assembly of Large RNA and DNA Genomes: Transmissible Gastroenteritis Virus Model

“…It will also be relatively easy to target TGEV to other species by simple replacements of the S glycoprotein gene (14,25,51). In contrast to arterivirus expression vectors, the coronavirus intergenic sequences rarely overlap upstream ORFs, simplifying the design and expression of foreign genes from downstream intergenic promoters (11,17,52). Several TGEV downstream ORFs also appear to encode luxury functions that can be deleted from the viral genome without affecting infectivity in vitro (18,29,56,57).…”

“…ORF1a encodes at least two viral proteases and several other nonstructural proteins, while ORF1b contains polymerase, helicase, and metal-binding motifs typical of an RNA polymerase (3,17,19). In the 3Ј-most ϳ9 kb of the TGEV genome, each of the downstream ORFs is preceded by a highly conserved intergenic sequence element, which directs the synthesis of each of the six or seven subgenomic RNAs (11,17,18,52). These subgenomic mRNAs are arranged in a nested set structure from the 3Ј end of the genome, and each contains a leader RNA sequence derived from the 5Ј end of the genome (26,29,42,43).…”

Characterization of the Coronavirus M Protein and Nucleocapsid Interaction in Infected Cells

“…M protein, the most abundant glycoprotein in the virus particle and in infected cells, is characterized as having three domains: a short N terminal ectodomain, a triple-spanning transmembrane domain, and a C-terminal endodomain (1). E protein is present only in minute amounts in infected cells and in the virus envelope (13,23,37,47,51), yet it is an essential protein for coronavirus envelope formation; coronavirus-like particles (VLPs) are assembled and released from cells that express both E and M proteins (4, 49). Furthermore, expression of E protein alone results in the production of membrane vesicles, which contain E protein (27).…”