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. 2005 May;79(10):6560–6564. doi: 10.1128/JVI.79.10.6560-6564.2005

FIG. 1.

FIG. 1.

Sequences, expression vectors, and tropism associated with prototypic MLV capsids. (A) Sequences of the capsid fragment used in the swaps are shown. Amino acid sequence alignment is shown for NB-tropic Friend MLV strain 57 and Moloney MLV (F-MLV and Mo-MLV, GenBank accession no. X02794 and J02255, respectively), B-tropic WNB5 MLV (GenBank accession no. K01190), N-tropic Akr-623 MLV (GenBank accession no. J01998), and the NT1 N-tropic Tennant strain of Friend MLV (this study; GenBank accession no. AY883167). NB, B, or N tropism is indicated in parentheses next to the MLV strain. The amino acid sequence encoded by the BamHI-BstXI 302-bp cassette (positions 1362 to 1664 in the Friend MLV clone 57 sequence), corresponding to amino acids 33 to 133 of the swapped capsid fragment, is shown. Positions corresponding to the EheI and XhoI internal restriction sites are also indicated. Distinctive residues including the NB determinant at positions 109 and 110 and the DIND motif (see text) at positions 92 to 95 are indicated. Dots represent residues identical to the Friend MLV 57 sequence. Each cassette was introduced in the CA gene of the pC57GP gag-pol expression vector from the NB-tropic strain 57 of Friend MLV. CMV, cytomegalovirus promoter; MA, matrix; CA, capsid; NC, nucleocapsid; pA, polyadenylation signal. (B) Evaluation of Fv1 and Ref1 MLV restrictions with parental CA cassettes. Virions were produced by transfecting human 293T cells with the vector shown in panel A and complementing vectors as described in the text. Viral supernatants were standardized on permissive Mus dunni Fv1−/− cells (dunni). Sensitivity to Fv1 and Ref1 restrictions was assayed on dunnin and dunnib cells stably expressing the Fv1n gene or the Fv1b gene, respectively, and on Ref1-positive human HT1080 cells. Cells were infected with serial dilutions of viral supernatant in the presence of Polybrene (8 μg/ml), and 2 days after infection, target cells were fixed and stained for alkaline phosphatase activity (20), and FFU per milliliter were counted. Mean viral titers ± standard errors of the means are shown and were calculated from at least three independent experiments using FFU values obtained in the linear portion of the titration curve.