Amino acid substitutions within the heptad repeat domain 1 of murine coronavirus spike protein restrict viral antigen spread in the central nervous system.
Targeted recombination was carried out to select mouse hepatitis viruses (MHVs) in a defined genetic background, containing an MHV-JHM spike gene encoding either three heptad repeat 1 (HR1) substitutions (Q1067H, Q1094H, and L1114R) or L1114R alone. The recombinant virus, which expresses spike with the three substitutions, was nonfusogenic at neutral pH. Its replication was significantly inhibited by lysosomotropic agents, and it was highly neuroattenuated in vivo. In contrast, the recombinant expressing spike with L1114R alone mediated cell-to-cell fusion at neutral pH and replicated efficiently despite the presence of lysosomotropic agents; however, it still caused only subclinical morbidity and no mortality in animals. Thus, both recombinant viruses were highly attenuated and expressed viral antigen which was restricted to the olfactory bulbs and was markedly absent from other regions of the brains at 5 days postinfection. These data demonstrate that amino acid substitutions, in particular L1114R, within HR1 of the JHM spike reduced the ability of MHV to spread in the central nervous system. Furthermore, the requirements for low pH for fusion and viral entry are not prerequisites for the highly attenuated phenotype.
Tsai
Jean C
JC
Department of Microbiology, University of Pennsylvania School of Medicine, Philadelphia, PA 19104-6076, USA.
de Groot
Linda
L
Pinon
Josefina D
JD
Iacono
Kathryn T
KT
Phillips
Joanna J
JJ
Seo
Su-hun
SH
Lavi
Ehud
E
Weiss
Susan R
SR
D019943:Amino Acid Substitution; D000818:Animals; D000956:Antigens, Viral / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D001921:Brain / Q000276:immunology / Q000378:metabolism*; D002459:Cell Fusion; D017934:Coronavirus / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*; D004274:DNA, Recombinant / Q000235:genetics; D008297:Male; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D009154:Mutation; D010641:Phenotype; D017434:Protein Structure, Tertiary; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D014779:Virus Replication
D000956:Antigens, Viral; D004274:DNA, Recombinant; D008562:Membrane Glycoproteins; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins