Identification of residues outside of the receptor binding domain that influence the infectivity and tropism of porcine endogenous retrovirus.
Identification of determinants of human tropism of porcine endogenous retrovirus (PERV) is critical to understanding the risk of transmission of PERV to recipients of porcine xenotransplantation products. Previously, we showed that a chimeric envelope cDNA encoding the 360 N-terminal residues of the human-tropic PERV envelope class A (PERV-A) SU and the 130 C-terminal residues of the pig-tropic PERV-C SU and all of TM (PERV-A/C) showed a 100-fold decrease in infectivity titer on human cells (M. Gemeniano, O. Mpanju, D. R. Salomon, M. V. Eiden, and C. A. Wilson, Virology 346:108-117, 2006). To identify residues important for human cell infection, we performed site-directed mutagenesis on each of the nine residues, singly or in combination, that distinguish the C-terminal region of PERV-C from PERV-A. Of the nine amino acids, two single-amino-acid substitutions, Q374R and I412V, restored the infectivity of human cells to the chimeric PERV-A/C to a titer equivalent to that of PERV-A. In contrast, PERV-A/C mutant envelope Q439P resulted in undetectable infection of human cells and an approximately 1,000-fold decrease in control pig cells. Mutation of K441R rescued mutants that carried Q439P, suggesting an incompatibility between the proline residue at this position and the presence of KK in the proteolytic cleavage signal. We confirmed this incompatibility with vectors carrying PERV-A envelope mutant R462K that were also rendered noninfectious. Finally, tropism of vectors carrying PERV-C envelope mutants with only four amino acid changes in the C terminus of PERV-C envelope, NHRQ436YNRP plus K441R, was shifted to one similar to that of PERV-A. Our results show an important and previously unrecognized role for infectivity and tropism for residues at the C terminus of SU.
Division of Cellular and Gene Therapies, Center for Biologics Evaluation and Research, FDA, 8800 Rockville Pike, Bethesda, MD 20892, USA.
Takele
T
Argaw
Mariel
M
Figueroa
Daniel R
DR
Salomon
Carolyn A
CA
Wilson
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D002460:Cell Line; D020077:Endogenous Retroviruses / Q000235:genetics / Q000502:physiology*; D015686:Gene Products, env / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013552:Swine; D053585:Virus Attachment
D015686:Gene Products, env; D011993:Recombinant Fusion Proteins