Targeted Recombination Demonstrates that the Spike Gene of Transmissible Gastroenteritis Coronavirus Is a Determinant of Its Enteric Tropism and Virulence

Abstract

Targeted recombination within the S (spike) gene of transmissible gastroenteritis coronavirus (TGEV) was promoted by passage of helper respiratory virus isolates in cells transfected with a TGEV-derived defective minigenome carrying the S gene from an enteric isolate. The minigenome was efficiently replicated in trans and packaged by the helper virus, leading to the formation of true recombinant and pseudorecombinant viruses containing the S proteins of both enteric and respiratory TGEV strains in their envelopes. The recombinants acquired an enteric tropism, and their analysis showed that they were generated by homologous recombination that implied a double crossover in the S gene resulting in replacement of most of the respiratory, attenuated strain S gene (nucleotides 96 to 3700) by the S gene of the enteric, virulent isolate. The recombinant virus was virulent and rapidly evolved in swine testis cells by the introduction of point mutations and in-phase codon deletions in a domain of the S gene (nucleotides 217 to 665) previously implicated in the tropism of TGEV. The helper virus, with an original respiratory tropism, was also found in the enteric tract, probably because pseudorecombinant viruses carrying the spike proteins from the respiratory strain and the enteric virus in their envelopes were formed. These results demonstrated that a change in the tropism and virulence of TGEV can be engineered by sequence changes in the S gene.

Comparative full-length genome sequence analysis of 14 SARS coronavirus isolates and common mutations associated with putative origins of infection

“…5 Differences in virulence of mice coronaviruses have also been linked to genetic variance in the S protein. 6,7 and the serological response in the host is typically raised against the S protein. 8 However, the S, M, and N mature proteins all contribute to generating the host immune response as seen in transmissible gastroenteritis coronavirus, 9 infectious bronchitis virus, 10,11 pig respiratory coronavirus, 12 and mouse hepatitis virus.…”

Altered pathogenicity, immunogenicity, tissue tropism and 3′-7kb region sequence of an avian infectious bronchitis coronavirus strain after serial passage in embryos

“…Using a reverse genetics system, the S glycoproteins for a group 2 coronavirus (MHV), a group 1 coronavirus (TGEV) and a group 3 coronavirus (IBV) were demonstrated to be involved in the tropism of these coronaviruses [26][27][28][29][30]68,70]. For the TGEV, several amino acid changes at the Nterminus of the S protein resulted in the loss of enteric tropism [29,30]. In this study, an amino acid substitution was found at residue 581 (Leu → Phe) between P70 and P110; however, loss of kidney tropism of CK/CH/LHLJ/04V occurred between P40 and P70.…”

“…The importance of S1 in determining cell and tissue tropism has been demonstrated for several coronaviruses, such as murine hepatitis virus [20][21][22][23][24][25][26][27][28], porcine transmissible gastroenteritis virus [29,30], and severe acute respiratory syndrome coronavirus [31]. In the case of IBV, the S1 subunit of the S protein determines the serotype of IBV and is responsible for viral attachment to cells.…”

Chimeric coronavirus-like particles carrying severe acute respiratory syndrome coronavirus (SCoV) S protein protect mice against challenge with SCoV

“…Coronavirus S protein is responsible for virus adsorption to susceptible cells through a specific virus-receptor interaction and induces membrane fusion between viral envelope and host cell membrane [5]. S protein is a main player for determining coronavirus tissue tropism, host specificity and viral pathogenicity [6][7][8][9][10][11][12]. Because most coronavirus neutralizing antibodies recognize S protein [1,13], it is not surprising that most of the current SCoV vaccine candidates are either the S protein subunit itself or those carrying S protein [14][15][16][17][18][19].…”