Frank Lilly

Genetic control of murine viral leukemogenesis.

Publisher Summary The question that whether the unit of malignancy is a gene, or an organism that grows in highly specialized protoplasmic environments, or a molecule that multiplies by virtue of integral chemical union with a molecule of protoplasm, remains unanswered. The question of whether leukemia in the mouse is caused by either genetic or viral mechanisms has taken on a new meaning with the recognition of host genetic constitution with the viral infection in leukemogenesis. The chapter covers the early studies of murine leukemia and the genetic control of friend virus susceptibility and naturally occurring leukemia viruses. It also discusses the h2 linked genetic control of leukemogenesis and the other genes responsible for leukemogenesis. Infection with ontogenic viruses is complex and malignancy is not an inevitable result of tumor virus infection; many specific events affect the interaction of leukemia virus with cells, leading to leukemogenesis.

Susceptibility to Two Strains of Friend Leukemia Virus in Mice

A mutant strain of the Friend leukemia virus is described. The parental virus strain, passaged through ICR/Ha mice, shows no or very little activity by the spleen-focus assay in BALB/c mice (by comparison with highly susceptible DBA/2 and ICR/Ha mice) and produces typical Friend disease in these mice only exceptionally; susceptibility to this strain of virus is recessive in the (C57BL/6 x DBA/2) F1. By contrast, the mutant virus strain is as active in BALB/c mice as in DBA/2 or ICR/Ha mice; susceptibility to the mutant virus strain is dominant in the (C57BL/6 x DBA/2) F1.

Selective incorporation of h-2 antigenic determinants into friend virus particles.

IN susceptible mice, Friend murine leukaemia virus (FV) produces fatal erythroleukaemia1 characterised by rapid onset of splenomegaly and viraemia in 4–5 d and leading to the death of the animal in as little as 3–4 weeks. In the replication of this enveloped, C-type RNA virus, assembly occurs at the surface of infected cells where the particles mature by budding2. We have examined the question of whether host cell surface glycoproteins are incorporated into the virion as the host cell membrane becomes part of the completed particle during the budding process. In particular, serologically identifiable alloantigens governed by the H–2K and H–2D genes of the major histocompatibility complex and found on cell surfaces have been sought in lysates of serum-derived FV.

B-tropic Friend virus: a host-range pseudotype of spleen focus-forming virus (SFFV).

Abstract The N-tropic strain of Friend virus (FV) complex includes two different infectious entities: defective SFFV and helper virus, the latter of which is responsible for the N-tropism of the complex. The N-tropic helper, highly infectious in homozygous Fv -1 n mice, is poorly infectious in Fv -1 b mice. B/T-L virus, a leukemogenic virus free of SFFV, includes a B-tropic helper virus which is much more infectious in Fv -1 b than in in Fv -1 n mice. N-tropic FV complex to which B/T-L virus has been added (i.e., SFFV with its associated N-tropic helper, plus B-tropic helper) induced FV disease in young Fv -1 b mice and gave rise to a strain of virus which could be passaged serially through Fv -1 b hosts. Analysis of the host range properties of the new virus strain indicated that the N-tropic helper component of FV complex had been rapidly eliminated during the first few passage generations of the virus, leaving a B-tropic pseudotype of SFFV, i.e., SFFV associated with B-tropic helper virus. Susceptibility to this B-tropic pseudotype was influenced in the expected manner by the Fv -1 and Fv -2 genes, but host range studies indicated that one or more previously unidentified genes also influenced the host response to this virus. Serial forced passage of the B-tropic FV through Fv -1 -restrictive hosts resulted in the recovery of an NB-tropic variant of the pseudotype.

Mechanisms of the H‐2 Effect on Viral Leukemogenesis

Evidence has been gathered which supports the notion that two distinct but interacting mechanisms, controlled by loci mapping within the H‐2 complex, influence Friend murine leukemia virus (FV) disease. One mechanism, controlled by a gene mapping in or close to H‐2D, influences the capacity of the H‐2D gene product to form molecular complexes with FV molecules in the plasma membrane of infected cells. Formation of a complex appears to provide a target antigen for syngeneic cytotoxic T‐lymphocytes, to cause co‐capping of FV and H‐2D antigens, to permit the selective inclusion of H‐2Db molecules into progeny Friend virions, to influence the long‐term maintenance of virus production in vitro and, in conjunction with the second mechanism, to stimulate the generation of cytotoxic T‐lymphocytes. This second mechanism is controlled by a gene in the H‐2K or H‐2I region, and, in the presence of an H‐2/FV molecular complex immunogen, influences the generation of H‐2 restricted cytotoxic T‐lymphocytes and the rate of rejection of syngeneic FV‐induced tumor cells.

Variations in viral gene expression in friend virus-transformed cell lines congenic with respect to the H-2 locus

Abstract The effect of H -2 genotype on the genetic expression of Friend oncovirus was studied using Friend virus-transformed cells differing genetically only at the H -2 locus. Seven lines and three sublines were examined with regard to expression of Friend virus and the murine leukemia virus envelope glycoprotein gp69/71, the structural proteins p30 and p15 (FMR), and reverse transcriptase. Our results confirm that H -2 b , H -2 g and H -2 kh cell lines were stable upon serial passage for production of infectious Friend virus, whereas the H -2 d and H -2 k cell lines ceased to release infectious virus after a relatively short time. Synthesis of viral proteins by these cell lines was independent of the synthesis of infectious virus in that some viral proteins continued to be synthesized by H -2 d and H -2 k cell lines in the absence of infectious virus. Reduction of viral proteins can be attributed to the loss of expression of individual viral genes. The temporal sequence of change suggests that the loss of protein expression may occur in a stepwise fashion; reverse transcriptase was reduced first, followed by p30 and p15, and gp69/71 last. The results confirm that p30 and p15 are coordinately expressed. In contrast, the expression of p30 and gp69/71 are noncoordinate, indicating that these proteins are regulated by independent genetic loci. Loss of viral genes did not occur, since cell lines which had previously ceased to synthesize viral proteins could be chemically induced to produce infectious NB tropic Friend viruses. These data suggest that for these H -2-congenic cell lines, there is an association of the H -2D b allele and the long-term production and release in vitro of both infectious virus and viral proteins.

Fv-1 Regulation of lymphoma development and of thymic, ecotropic and xenotropic muLV expression in mice of the AKR/J × RF/J Cross

The incidence of spontaneous thymic lymphoma has been studied in crosses between AKR/J and RF/J mice. AKR mice develop a high incidence of this disease. RF mice transmit a marked resistance to development of the disease to F1 hybrid mice of the AKR x RF cross. This resistance is associated with a reduction of endogenous ecotropic and xenotropic MuLV expression in the prelymphomatous thymus. The RF gene governing the coordinate suppression of these three phenotypes has been mapped to the Fv-1 locus. These results indicate that the particular Fv-1 allele of AKR mice provides a permissive genetic background for endogenous ecotropic and xenotropic MuLV expression and that these viral activities may be etiologically involved in the development of spontaneous thymic lymphoma in the mouse.

Identification of an epitope encoded in the env gene of friend murine leukemia virus recognized by anti-friend virus cytotoxic T lymphocytes

We have previously shown that strong epitopes recognized by anti-Friend virus (FV) cytotoxic T lymphocytes (CTL) in H-2b mice are encoded in both the env and gag/pol regions of the helper friend leukemia virus genome. Two approaches have been used to identify these epitopes. At the nucleic acid level, we have constructed env genes with either of two in-frame deletions: pKR2, an env gene with a 681-bp deletion in the gp70 region and inserted into the pSV2-gpt-1 expression vector; and pKR1, an env gene with an 81-bp deletion in the p15E region and inserted into pSV2-gpt-1. Cell clones were established by transfecting Fisher rat embryo cells with pDb (the H-2Db restriction element), pNEO (for G418 selection) and either pKR1 or pKR2. Db and env gene expression was monitored by immunoprecipitation with polyclonal antibodies or by detection of viral RNA on Northern blots. Expressor cell clones were tested for susceptibility to lysis by polyclonal anti-FV/Db CTL in 51Cr-release assays. Whereas cells expressing pKR1 were lysed to the same extent as cells expressing the intact env gene, cells expressing pKR2 were resistant to lysis, suggesting that all detectable env epitopes are encoded within the 681-bp deletion. Polypeptides representing the two most likely candidate epitopes encoded in this segment were synthesized and tested for their abilities to sensitize FRE cells expressing Db alone for lysis by the CTL. One 17-mer polypeptide, AGTGDRLLNLVQGAYQA [corrected], functioned as a strong CTL epitope in this assay, but the other 18-mer polypeptide was inactive. Studies of the role of this epitope in the immune response to candidate viral vaccines are in progress.

Detection of h-2db antigen on osmotic shock-treated friend leukemia virus particles.

Antisera specific for either H-2Kb, H-2Db, H-2Kk or H-2Dk antigenic determinants were examined for their capacity to neutralize Friend virus (FV) collected from the serum of infectedH-2b/H-2k heterozygous mice. Neutralizing activity was detected (1) only withanti-H-2Db antisera, (2) only when the surface of virus particles had been mildly deranged by osmotic shock treatment and (3) only in the assay for the defective spleen focus-forming virus component of FV.

The effect of immunosuppression on oncogenesis by murine sarcoma virus.

Summary In adult CBA/Wh mice the incidence and mean latency periods of tumors appearing following infection with MSV were dependent upon the dose of virus inoculated and the immunological state of the host. Animals of reduced immunological competence had a 10-fold higher susceptibility to tumor induction as well as a decreased latency period to tumor appearance compared with controls. Moreover, whereas these tumors regressed in intact adult mice, they tended to grow progressively in immunosup-pressed hosts. These results indicate that the immunological competence of the host is a prime determinant of susceptibility to the oncogenic action of murine sarcoma virus.