title: Single amino acid substitution (G42E) in the receptor binding domain of mouse mammary tumour virus envelope protein facilitates infection of non-murine cells in a transferrin receptor 1-independent manner.
abstract: BACKGROUND
  Mouse mammary tumour virus (MMTV) is a betaretrovirus that infects rodent cells and uses mouse tranferrin receptor 1 (TfR1) for cell entry. Several MMTV strains have been shown to productively infect, in addition to murine cells, various heterologous cell lines including those of human origin, albeit less efficiently than murine cells. Furthermore, there have been reports that the continued passage of MMTV in heterologous cell lines gives rise to novel variants that are able to infect naive non-murine cells with higher efficiency than the parental virus.
  
  
  RESULTS
  We show that MMTV(C3H), like other MMTV strains, that had undergone a number of replication cycles in non-murine cells displayed an increased replication kinetic, as compared to parental virus, when applied on naive human cells. Sequence analysis of several replication kinetic variants and the parental virus, together with calculation of the ratio of non-synonymous to synonymous mutations at individual codons, revealed that several regions within the viral genome were under strong positive selection pressure during viral replication in human cells. The mutation responsible, at least in part, for the phenotypic change was subsequently mapped to the segment of env encoding the receptor binding site (F₄₀HGFR₄₄). Introduction of the identified mutation, leading to single amino acid substitution (G42E), into egfp-containing recombinant MMTV virions enhanced their ability to bind to and infect human cells. Interestingly, neither the replication kinetic mutant nor the parental virus required human TfR1 for infection. Knock-out of TFR1 gene from the human genome did not decrease the susceptibility of Hs578T cells to virus infection. Furthermore, the expression of human TfR1, in contrast to mouse TfR1, did not enhance the susceptibility of MMTV-resistant Chinese hamster ovary cells. Thus, human TfR1 is dispensable for infection and another cell surface molecule mediates the MMTV entry into human cells.
  
  
  CONCLUSIONS
  Taken together, our data explain the mechanism enabling MMTV to form 'host-range variants' in non-murine cells that has been known for a long time, the basis of which remained obscure. Our findings may expand our understanding of how viruses gain capability to cross species-specific barriers to infect new hosts.
authors.affiliation: Institute of Virology, University of Veterinary Medicine, Veterinaerplatz 1, 1210, Vienna, Austria. Constantine.Konstantoulas@vetmeduni.ac.at.
authors.forename: Constantine James
authors.initials: CJ
authors.lastname: Konstantoulas
authors.affiliation: Institute of Virology, University of Veterinary Medicine, Veterinaerplatz 1, 1210, Vienna, Austria. benjamin.lamp@vetmeduni.ac.at.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Lamp
authors.affiliation: Institute of Virology, University of Veterinary Medicine, Veterinaerplatz 1, 1210, Vienna, Austria. Till.Ruemenapf@vetmeduni.ac.at.
authors.forename: Tillman Hans
authors.initials: TH
authors.lastname: Rumenapf
authors.affiliation: Institute of Virology, University of Veterinary Medicine, Veterinaerplatz 1, 1210, Vienna, Austria. Stanislav.Indik@vetmeduni.ac.at.
authors.forename: Stanislav
authors.initials: S
authors.lastname: Indik
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D002460:Cell Line; D058507:Host Specificity; D006801:Humans; D008324:Mammary Tumor Virus, Mouse / Q000502:physiology*; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism*; D020125:Mutation, Missense; D011990:Receptors, Transferrin / Q000378:metabolism*; D012692:Serial Passage; D014759:Viral Envelope Proteins / Q000235:genetics / Q000378:metabolism*; D053586:Virus Internalization; D014779:Virus Replication
chemical_list: D050505:Mutant Proteins; D011990:Receptors, Transferrin; D014759:Viral Envelope Proteins