Variations in properties of virus released from morphologically different cell lines transformed in vitro by Friend leukemia virus.

Abstract

The properties of the virus synthesized by each of three morphologically different cell lines originating from DBA/2J fetal liver cells transformed by the anemic strain of Friend leukemia virus in vitro were analyzed. The cells of line C-1 are malignant in syngeneic DBA/2 mice, grow in suspension, and are erythroid in origin. Cells of lines G-2 and G-3 are adherent, are epithelial in appearance, and produce no tumors in DBA/2J mice. Higher reverse transcriptase activity was detected in the culture fluid of lines G-2 and G-3, although virus from G-1 cells was more leukemogenic. Differences were also found in the virion density and size of the viral genome. RNA from the virions produced by G-2 and G-3 cells sedimented at 75 S in a sucrose gradient; virion RNA from C-1 cells sedimented at 60 S. However, when subjected to polyacrylamide gel electrophoresis, all three virus strains showed identical RNA subunits with an estimated molecular weight of 2.6 X 106. Analysis of virion proteins by slab gel electrophoresis showed differences in envelope protein (gp7l) components but not in the major core protein (p30). The properties of these viruses are stable and remain unchanged after passage in 3T3 cells.The anemic strain of Friend leukemia virus (FLV-A) is a competent virus with the ability to induce erythroleukemia in the absence of spleen focus-forming virus (SFFV) (1). The primary target cell for FLV is most likely an erythroid precursor (2). That infection may also affect other cell types (3) was supported by the finding that permanent lines of morphologically. different cells originated from cultures of fetal liver transformed by FLV-A in vitro. Three of these lines have been characterized (4). The cells of line G-I are immature hematological cells and resemble the erythroleukemia lines established from the spleens of FLV-infected mice (5). They grow in suspension and can be induced to differentiate along the erythroid pathway. The cells of lines G-2 and G-3 are adherent and epithelial. All three lines are chronically infected with virus.In view of the findings that greater levels of virus were produced by the adherent lines and that this virus was less leukemogenic than that produced by the suspension lines, it was important to compare the molecular properties of the virus of each of the three transformed cell lines. The virus produced by the adherent lines was found to differ from that of the suspension line in the virion particle density, sedimentation coefficient of high molecular weight (HMW) RNA, and viral envelope protein components. However, all of the virus strains have identical RNA subunits.MATERIALS AND METHODS Cells and Viruses. Establishment, growth conditions, and characterization of the FLV-A-transformed lines of G-1, G-2, and G-3 (4) as well as of the prototype Friend erythroleukemia cell line 745A (5) and its subclone 5-86 (6, Virus Replication. Virus released into the culture fluid was assayed for syncytia formation on XC cells (9). Generally, leukemogenicity was assayed in 6-to 8-week-...

Rat cells infected with anemia-inducing Friend leukemia virus contain integrated replication-competent but not defective proviral genomes.

“…Thus, the role of the 5.7-kb Kpn I fragment found in multiple copies in the FLV-A-transformed erythroleukemic cell lines SQ-A, G-1, and 5-86 remains unknown. It does not appear to be associated with pathogenicity, since the virus from G-1 and 5-86 cultures have little or no leukemogenic activity (15).…”

“…Cells were maintained in Glasgow minimum essential medium (GIBCO) supplemented with 10% fetal calf serum, penicillin, and streptomycin. Cultures of 3Y1 and NIH 3T3 cells were infected with FLV-A prepared from leukemic spleen filtrates of adult DBA/2 mice as described (15).…”

“…Several wild-type FLV-A-transformed cell lines produce virus that differ in such properties as particle density, the sedimentation coefficient of virion RNA, and viral-envelope proteins and have little or no leukemogenic activity (15). However, the more recently isolated cell line SQ-A continues to produce leukemogenic virus (16).…”

Characterization of leukemogenic virus produced by a new line of Friend erythroleukemia virus-transformed cells.

“…The amount of FLV released by the chronically infected cells was measured by assay for syncytia on XC cells (6) and by assaying the reverse transcriptase activity released into the medium. The enzymatic activity was quantitated by incorporation of [3H]thymidine monophosphate into acid-soluble material using poly(rA-dT) as template primer under the conditions as described (3,7). To test for leukemogenic activity, 0.2-0.5 ml of Millipore 0.45-,m filtered, 4-day supernatant fluid was inoculated intraperitoneally into Swiss (Taconic Farms, Germantown, NY) or DBA/2J (The Jackson Laboratory) 8-to 10-week-old mice.…”