Leukemia Viruses Associated with Mouse Myeloma Cells

Abstract

Abstract. Myeloma cells derived from BALB/c and C3H mice show evidence of infection by a murine leukemia virus. The immunoglobulin-producing myelomas secrete an RNA-containing virus with a density of 1.20 to 1.22 gm/cm3. RNA with a sedimentation coefficient of 74 S in 0.1 M sodium chloride has been isolated from secreted virus particles and has a base composition similar to that found for other murine leukemia virus RNA. An intracellular virus particle has been partially purified and has a density of 1.29 to 1.32 gm/cm3. Both extracellular and intracellular virus particles contain the leukemia virus group-specific antigen.Ultrastructural characteristics of viruslike particles in the cytoplasm of mouse plasma cell tumors have been described.1-' Here we identify the extracellular and intracellular particles from cultures of myeloma cells secreting immunoglobulin, and from spontaneous variants of these cell lines which secrete only the heavy or light chains of immunoglobulins, as viruses related to the murine leukemia viruses.Materials and Methods. Cell lines: C1 cells were derived from a myeloma of a C3H mouse; the other tumor lines were derived from oil-induced myelomas of BALB/c mice (see Table 1). Cell lines were cloned in the spleen or in agar,4 and grown as suspensions in fortified Eagle's medium'0 supplemented with 10 to 20% horse serum. All experiments were performed on exponentially growing cultures with doubling times of 18 to 24 hr.Virus purification: (a) Extracellular virus: Cells and debris were removed from the medium by two centrifugation steps (150 g for 5 min and 12,000 g for 10 min). The medium was layered over 2 ml 15% potassium tartrate containing 0.1 M Tris-HCl buffer (pH 8.0) and centrifuged in a Spinco SW25.3 rotor at 25,000 rpm for 60 min. The pellet was resuspended in 0.1 M Tris-HCl buffer (pH 8.0) and layered over a 4.8-ml gradient of 15 to 60% potassium tartrate containing 0.1 1I1 Tris-HCI buffer (pH 8.0)," and centrifuged in a Spinco SW65 rotor at 35,000 rpm for 60 min.(b) Intracellular virus: Cells were washed twice in NET buffer (0.1 M NaCl, 0.05 M Tris-HCl (pH 7.4) 0.001 ill EDTA), collected by low speed centrifugation, resuspended in NET buffer (approx. 106 cells/ml), and stored at -15'C until used.After thawing, Nonidet P40 was added to cell suspensions to a final volume of 1%, the solution pipetted vigorously to ensure lysis of all cells, and then centrifuged at 12,000 g for 10 min. The supernatant was layered over 2 ml 15% sucrose containing NET and centrifuged for 60 min in a SW65 rotor at 30,000 rpm. The pellet was resuspended in 0.1 M Tris-HCl buffer (pH 8.0) and analyzed by tartrate density-gradient centrifugation.CsCl-equilibrium sedimentation: Virus solutions were layered over 5 ml previously formed CsCI gradients (1.08-1.43 gm/cm3) and centrifuged for 20 hr at 15'C in a Spinco SW65 rotor at 50,000 rpm.

Some Structural and Antigenic Properties of Intracisternal A Particles Occurring in Mouse Tumors

“…In experiments to be described fully in another context, MuLV group specific antigen was demonstrable by CF assay in RPC-20 membrane fractions but failed to fractionate with the A-particles released by either the Triton-shear procedure or sonic vibration. Our results thus far do not support the suggestion that intracisternal A-particles in BALB/c myeloma cells represent intracellular or incomplete forms of MuLV (27,28).…”

A Factor That Can be Used to Regulate an In Vitro Primary Immune Response

“…Mouse myelomas and lymphomas were derived from oil-induced tumors of BALB/c mice (8). All myeloma and lymphoma cells used here were infected with Gross-type leukemia viruses (9).…”

“…The factor found in supernatants from MI1, or JLS-V5 cell cultures is not secreted by all cell lines infected with murine leukemia viruses. A number of plasmacytomas known to be infected with Gross-type leukemia viruses (9) and 3T3 cells infected with Rauscher leukemia virus do not secrete the factor. Uninfected spleen cells, however, do secrete the factor (Table 2).…”

“…Erythroleukemic -cells infected with Friend leukemia virus have been induced to differentiate along the erythroid pathway (17). Myelomas derived from BALB/c mice by oil-induction (18,19) are bone marrow-derived lymphocytes infected with Gross-type leukemia viruses (9) that continue to synthesize antibody 'or maintain their differentiated state in the absence of antigenic and accessory cell stimulation. If Mb-1, ML-2, and JLS-V5 cells have been derived from cells whose physiological function is to secrete factor, then infection with leukemia viruses has resulted in a specific cell, which maintains a normal function.…”

Canine Systemic Lupus Erythematosus. TRANSMISSION OF SEROLOGIC ABNORMALITIES BY CELL-FREE FILTRATES

“…Tumor cells and cell-free filtrates from two transplantable lymphoid tumors of mice (MOPC 315 [11]) and GVH (12) that were not in-(luced by the canine filtrate, but which are known to contain murine leukemia virus, failed to elicit ANA production after administration to normal newborn puppies. to N-DNA-can be induced in appropriate recipients by cell-free filtrates of tissues from dogs with ANA and positive LE cell tests.…”