Tolerance to mutations in the foot-and-mouth disease virus integrin-binding RGD region is different in cultured cells and in vivo and depends on the capsid sequence context.
Engineered RNAs carrying substitutions in the integrin receptor-binding Arg-Gly-Asp (RGD) region of foot-and-mouth disease virus (FMDV) were constructed (aa 141-147 of VP1 capsid protein) and their infectivity was assayed in cultured cells and suckling mice. The effect of these changes was studied in the capsid proteins of two FMDVs, C-S8c1, which enters cells through integrins, and 213hs(-), a derivative highly adapted to cell culture whose ability to infect cells using the glycosaminoglycan heparan sulfate (HS) as receptor, acquired by multiple passage on BHK-21 cells, has been abolished. The capsid sequence context determined infectivity in cultured cells and directed the selection of additional replacements in structural proteins. Interestingly, a viral population derived from a C-S8c1/L144A mutant, carrying only three substitutions in the capsid, was able to expand tropism to wild-type (wt) and mutant (mt) glycosaminoglycan-deficient CHO cells. In contrast, the 213hs(-) capsid tolerated all substitutions analysed with no additional mutations, and the viruses recovered maintained the ability of the 213hs(-) parental virus to infect wt and mt CHO cells. Viruses derived from C-S8c1 with atypical RGD regions were virulent and transmissible for mice with no other changes in the capsid. Substitution of Asp143 for Ala in the C-S8c1 capsid eliminated infectivity in cultured cells and mice. Co-inoculation with a neutralizing monoclonal antibody directed against the type C FMDV RGD region abolished infectivity of C-S8c1 virus on suckling mice, suggesting that FMDV can infect mice using integrins. Sequence requirements imposed for viral entry in vitro and in vivo are discussed.
Gutiérrez-Rivas
Mónica
M
Centro de Investigación en Sanidad Animal, INIA, Valdeolmos, 28130 Madrid, Spain.|Centro de Biología Molecular Severo Ochoa (CSIC-UAM), Cantoblanco, 28049 Madrid, Spain.
Pulido
Miguel Rodríguez
MR
Centro de Investigación en Sanidad Animal, INIA, Valdeolmos, 28130 Madrid, Spain.|Centro de Biología Molecular Severo Ochoa (CSIC-UAM), Cantoblanco, 28049 Madrid, Spain.
Baranowski
Eric
E
Université de Toulouse, ENVT, UMR 1225, F-31076 Toulouse, France.|INRA, UMR 1225, F-31076 Toulouse, France.
Sobrino
Francisco
F
Centro de Investigación en Sanidad Animal, INIA, Valdeolmos, 28130 Madrid, Spain.|Centro de Biología Molecular Severo Ochoa (CSIC-UAM), Cantoblanco, 28049 Madrid, Spain.
Sáiz
Margarita
M
Centro de Investigación en Sanidad Animal, INIA, Valdeolmos, 28130 Madrid, Spain.|Centro de Biología Molecular Severo Ochoa (CSIC-UAM), Cantoblanco, 28049 Madrid, Spain.
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D000833:Animals, Suckling; D001665:Binding Sites / Q000235:genetics; D016466:CHO Cells; D036022:Capsid Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D005536:Foot-and-Mouth Disease / Q000473:pathology* / Q000821:virology; D029544:Foot-and-Mouth Disease Virus / Q000235:genetics / Q000378:metabolism / Q000472:pathogenicity*; D039302:Integrin alphaVbeta3 / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D009842:Oligopeptides / Q000737:chemistry / Q000235:genetics*; D011991:Receptors, Virus / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014774:Virulence
D036022:Capsid Proteins; D039302:Integrin alphaVbeta3; D009842:Oligopeptides; D011991:Receptors, Virus; C442501:VP1 protein, Foot-and-mouth disease virus; C047981:arginyl-glycyl-aspartic acid