Marilyn R. Lander

Cellular origin and role of mink cell focus-forming viruses in murine thymic lymphomas

Embryo DNA of AKR mice contains several copies of intact mink cell focus-forming (MCF)-like env gene sequences with a proviral structure. In spontaneous thymic tumours, these MCF-like env sequences have regularly recombined with a specific region of the 3′ end of ecotropic env. This specific recombination is probably a critical event in the pathogenesis of spontaneous AKR tumours.

Noninfectious ARK mouse embryo cell lines in which each cell has the capacity to be activated to produce infectious murine leukemia virus

Abstract When cells of AKR mouse embryos were grown in tissue culture, only one cell in 250,000 to one in 12,000,000 produced murine leukemia virus within the first few days in culture. By 2.5 weeks in culture, 12–100 times this many cells had spontaneously begun to produce virus. By planting cultures with small numbers of AKR embryo cells, it was possible to obtain two cell lines which contained no detectable virus-producing cells for more than 60 serial transfers, despite exhaustive tests for virus and virus products. However, these lines and all 10 clonally derived sublines have the capacity to produce virus, either spontaneously or after certain experimental manipulations, including X-ray or ultraviolet irradiation, and transformation by SV40 virus. Activation of virus appears to be a very low-frequency event. These findings indicate that the majority, and probably all, of the cells in the AKR cell lines carry the full viral genome in an unexpressed form, and by extrapolation suggest that this is true of all AKR cells.

Origin of mink cytopathic focus-forming (MCF) viruses:Comparison with ecotropic and xenotropic murine leukemia virus genomes

Abstract Restriction endonuclease maps have been developed for the viral DNAs from nine xenotropic and eleven MCF murine leukemia viruses (MuLV) isolated from AKR and other mouse strains. In contrast to the highly related nature of ecotropic viral DNAs isolated from inbred mice and from M. m. molossinus , each xenotropic and MCF viral DNA was unique. Xenotropic MuLV DNAs could be divided into two classes, which correlated with previously reported serological and biochemical data; one xenotropic MuLV isolated from an AKR mouse showed features of both classes. Ecotropic viral DNA hybridized poorly or not at all to a 1.2 kbp segment of xenotropic viral DNA located in env 6.3–7.5 kbp from the left end of the viral DNAs. All MCF viral DNAs contained noneco-tropic sequences in a portion of the env gene region, but some MCF viruses were composed principally of nonecotropic sequences. The nonecotropic regions of the MCF viral DNAs were related to xenotropic MuLV DNA, but many MCF viral DNAs contained sequences not found either in xenotropic or ecotropic MuLV DNA. It was concluded that these MCF viruses probably arose via recombination between ecotropic MuLV and endogenous MuLV DNA sequences; sequences of recombination include a portion of env, but need not be limited to this region. The polytropic host range of MCF viruses may represent an endogenous viral function.

Accelerated Appearance of Multiple B Cell Lymphoma Types in NFS/N Mice Congenic for Ecotropic Murine Leukemia Viruses

Spontaneous lymphomas occur at high frequency in NFS.V+ mice, strains congenic for ecotropic murine leukemia virus (MuLV) proviral genes and expressing virus at high titer. In the present study, a total of 703 NFS.V+ lymphomas were studied by histopathology, immunophenotypic analysis, immunoglobulin heavy chain or T cell receptor β chain rearrangements, and somatic ecotropic MuLV integrations; 90% of the lymphomas tested were of B cell lineage. Low-grade tumors included small lymphocytic, follicular, and splenic marginal zone lymphomas, while high-grade tumors comprised diffuse large-cell (centroblastic and immunoblastic types), splenic marginal zone, and lymphoblastic lymphomas. Comparison of mice of similar genetic background except for presence (NFS.V+) or absence (NFS.V−) of functional ecotropic MuLV genomes showed that NFS.V− clonal lymphomas developed at about one-half the rate of those occurring in NFS.V+ mice, and most were low-grade B cell lymphomas with extended latent periods. In NFS.V+ mice, clonal outgrowth, defined by Ig gene rearrangements, was associated with acquisition of somatic ecotropic proviral integrations, suggesting that, although generation of B cell clones can be virus independent, ecotropic virus may act to increase the rate of generation of clones and speed their evolution to lymphoma. The mechanism remains undefined, because only rare rearrangements were detected in several cellular loci previously associated with MuLV insertional mutagenesis.

Molecular phylogeny of Fv1

Abstract. Alleles at the Fv1 gene of inbred mice confer resistance to infection and spread of vertically or horizontally transmitted murine leukemia viruses (MuLV). The nucleotide sequence of Fv1 bears similarity to the gag of a human endogenous retrovirus, HERV-L, but is more closely related to the gag-coding sequence of a newly described class of HERV-L-related mouse endogenous retroviruses designated MuERV-L. Both observations suggest an origin of Fv1 from endogenous gag sequences. The molecular definition of Fv1 provided an opportunity to determine the phylogeny of the gene among wild mice and its relation to MuERV-L. PCR primers, chosen to include most of the coding region of Fv1 for both the n and b alleles, were used to amplify sequences from animals of the genus Mus, which were then sequenced. Closely related products were obtained from almost all animals examined that evolved after the separation from Rattus, in which the homologous gene was shown to be absent. A phylogenetic tree generated with Fv1 sequence data differs noticeably from that developed with sequence data from other genes. In addition, non-synonymous changes were found to be present twice as frequently as synonymous changes, a fact that departs from the standard behavior of a structural gene. These observations suggest that the Fv1 gene may have been subjected to possible horizontal transfers as well as to positive Darwinian selection.

Amplification and rearrangement of onc genes in mammalian species

Related eukaryotic species usually contain the same number of copies per cell of a given ‘unique sequence’ gene. In the few described exceptions, such as the preproinsulin1,2 and globin genes3–7, one or two additional copies per cell have been found in several related species, suggesting that germ-line amplification occurred millions of years ago in a common progenitor of these species. The transforming (onc) genes of retroviruses are a group of evolutionarily conserved genes for which at least 10 distinct members have been described (for a review see ref. 8). Sequences related to each onc have been identified in all vertebrate species tested, usually as one copy per haploid genome, although the rat has at least two different genes homologous to the onc of the rat-derived Harvey murine sarcoma virus (Ha-MuSV)9. In screening genomic DNA from several rodent species for sequences related to the onc of Ha-MuSV and to the closely related (but distinguishable) onc of Kirsten (Ki) MuSV10, we have now found evidence for relatively recent amplification of these genes. We report here that Mus pahari apparently contains at least 10 copies of Ha-MuSV-type onc, whereas most other Mus species contain only one or two copies. Similarly, Chinese hamsters (Cricetulus iseus) have about six copies of the Ki-MuSV-type onc compared with only one copy in Syrian hamsters (Mesocricetus auratus).

Lymphomas and High-Level Expression of Murine Leukemia Viruses in CFW Mice

ABSTRACT Historically, Swiss Webster mice of the CFW subline, both inbred and random-bred stocks, have been considered to have a low spontaneous occurrence of hematopoietic system tumors, and previous reports of infectious expression of murine leukemia viruses (MuLVs) have been rare and unremarkable. In marked contrast, in the present study of CFW mice from one source observed by two laboratories over a 2-year period, nearly 60% developed tumors, 85% of which were lymphomas, the majority of B-cell origin. All tumors tested expressed ecotropic MuLVs, and most expressed mink cell focus-inducing (MCF) MuLVs. Among normal mice of weanling to advanced age, over one-half were positive for ecotropic virus in tail or lymphoid tissues, and MCF virus was frequently present in lymphoid tissue, less often in tail. Patterns of ecotropic proviral integration indicated that natural infection occurred by both genetic and exogenous routes. Lymphomas were induced in NIH Swiss mice infected as neonates with tissue culture-propagated MuLVs isolated from normal and tumor tissue of CFW mice.

Restriction endonuclease mapping of ecotropic murine leukemia viral dnas: size and sequence heterogeneity of the long terminal repeat.

Abstract A physical map for 12 restriction endonucleases has been derived for the unintegrated linear viral DNA of 11B3 murine leukemia virus (MuLV), which is the endogenous ecotropic XC plaque-forming virus released from the cell line from which integrated MuLV molecular clones 614 and 623 were isolated. This virus had originally been isolated from a mouse carrying the Akv-1 locus. The 11B3 MuLV DNA map was compared with the maps derived for the DNAs of several other endogenous ecotropic MuLVs isolated from inbred mouse strains and from the Asian feral mouse Mus musculus molossinus . In general, their maps were highly related, indicating a common origin for most of their sequences. However, 11B3 MuLV DNA differed from the other MuLV DNAs in that its long terminal repeat (LTR) was about 50 by longer than that of the other MuLVs and it lacked a Hind III site which was located 3 kbp from the 5′ end in the other MuLVs. These two differences in 11B3 MuLV probably arose after virus activation in the mouse, perhaps by recombination with other endogenous MuLV sequences. In the endogenous ecotropic MuL V DNAs (including 11B3 MuLV), the U3 region of the LTR differed from this region in Moloney MuLV for each of the five restriction endonaclease sites which were tested. We conclude that the LTR of murine C-type viruses may contain size and sequence heterogeneity.

Expression of cyclin D1 in mouse B cell lymphomas of different histologic types and differentiation stages

The G1 cyclin, cyclin D1, has been implicated in the development of human and mouse tumors. Here we describe immunohistochemical analyses of cyclin D1 for a large panel of mouse B cell tumors. In addition, we characterize cyclin D1 expression in a series of cultured cell lines that represent transformed B cells at different stages of development. Immunohistochemical analysis showed that for low-grade lymphomas, cyclin D1 was expressed by 83% of centroblastic centrocytic (CBCC) and 14% of small lymphocytic lymphomas (SLL). For high-grade tumors, 28% of B lymphoblastic and 23% of centroblastic tumors expressed cyclin D1, while all immunoblastic lymphomas were negative. Studies of RNA and protein prepared from cultured B lineage tumors showed that cyclin D1 was expressed by all pre-B and most B cell tumors but not by cell lines representative of late B cell differentiation or by plasma cells. Expression of cyclin D1 in the lymphomas was not associated with alterations in the genomic structure of the Fis-1 (Bcl-1) common proviral integration site or cyclin D1 itself or with cell growth activity as assessed by expression of proliferating cell nuclear antigen (PCNA).

Regulation of endogenous type C viruses: Evidence for transcriptional control of AKR viral expression

Abstract Nuclear and cytoplasmic RNAs isolated from AKR virus producer and virus-negative cell lines, as well as in vitro transcripts of chromatin isolated from these cells, were analyzed by nucleic acid hybridization using a 3 H-labeled DNA complementary to the genome of the endogenous AKR ecotropic murine leukemia virus. Our results suggest that regulation of expression of the endogenous virogene sequences resides in part at the transcriptional level.