Avian Myelocytomatosis Virus Strain MC29 Research Articles

Effects of chicken anaemia virus on experimental leukosis, induced by avian myelocytomatosis virus Mc29

The effects of concomitant infection with chicken anaemia virus (CAV) on the incidence, clinical manifestation and mortality from leukosis, induced by the avian myelocytomatosis virus strain Mc29 were studied. Experimental one-day-old 15 I line White Leghorn chickens were inoculated simultaneously with Mc29 and CAV or with Mc29 alone and observed daily for clinical signs and mortality. Both groups of chickens inoculated with Mc29 virus strain alone or in combination with CAV deve­loped tumours and died within 57 days. Necropsy has been performed on all dead birds following the standard protocol. Organ samples from thymuses, spleens, bone marrow, and livers were collected and histopathologically investigated. Neoplasms detected included myelocytomas, nephroblastomas and hepatocellular carcinomas. In addition, 50% of the CAV/Mc29-inoculated chickens developed epithelial type thymomas. However, no such lesions were found in chickens infected with Mc29 alone. No significant differences in the clinical course of leukosis between the two experimental groups of chickens were observed. The results indicated that CAV infection did not affect substantially the incidence and mortality from avian leukosis, induced by myelocytomatosis virus strain Mc29, but contributed to greater variety of the induced tumours.

Myc products induce the expression of catecholaminergic traits in quail neural crest-derived cells.

The avian myelocytomatosis virus strain MC29 v-myc oncogene transforms a wide panel of avian cells in vitro and either blocks or maintains differentiation, depending on the cell type. In the present work, we have investigated the effect of this oncogene on the differentiation of early embryonic cells, neural crest cells, grown in vitro. We report that the MC29 v-myc gene product induces a strong cellular proliferation of 2-day quail neural crest with the appearance of catecholaminergic traits. Other v-myc as well as the c-myc gene products also trigger this phenotype. Retroviruses carrying some other oncogenes do not elicit this phenotypic expression, although they activate cell multiplication. Thus, our results indicate that myc gene products induce (directly or indirectly) a differentiated phenotype in a subpopulation of neural crest cells.

Heart tumors specifically induced in young avian embryos by the v-myc oncogene.

To determine if expression of the v-myc oncogene had any effect during ontogeny, we injected avian myelocytomatosis virus strain MC29 into avian embryos at various stages of development. The injection of MC29 at embryonic day 2 (E2) or 3 (E3) caused, about 10 days later, rhabdomyosarcomas of the heart and, in some cases, skin muscle hypertrophy. When the injection was performed at E4 or E5, the number of heart tumors declined, whereas the number of skin muscle tumors increased significantly. The p110gag-myc protein was found in all tumors analyzed. When the virus was injected intravenously into E10 embryos, no tumors appeared during embryonic life, in striking contrast to the results obtained from injections at earlier stages. The monoclonal antibody 13F4, which is specific for the myogenic lineage, bound strongly to tumoral heart tissue, whereas it bound weakly to normal cardiac cells. Comparison of the peaks of tumor incidence in relationship to the timing of injection suggests that the v-myc product could interfere in vivo with an early step of the muscle lineage differentiation program. In addition, we show that the p58c-myc protein, which is supposed to play an important role in the control of cell proliferation, is only faintly detected in the heart of normal E3 embryos, in contrast to limb and tail buds, which readily express detectable levels of p58c-myc.

Avian myelocytomatosis virus immortalizes differentiated quail chondrocytes.

Quail embryo chondrocytes in culture display two morphological phenotypes: polygonal epithelial-like and floating cells. Both cell populations synthesize cartilage extracellular matrix proteins (type II collagen and specific proteoglycans), whereas type X collagen, which appears to be a marker of later stages of chondrocyte differentiation, is expressed only by the epithelial-like cells. Avian myelocytomatosis virus strain MC29 does not induce morphological transformation in quail embryo chondrocytes but stimulates these cells to proliferate with a progressively reduced doubling time. MC29-infected chondrocytes can be established in culture as a continuous cell line, whereas control (uninfected) cultures only survive a few months. Rapidly dividing MC29-infected chondrocytes still express type II collagen and cartilage proteoglycans but do not synthesize type X collagen.

Virus gene expression in rat cells transformed by avian myelocytomatosis virus strain MC29 and avian erythroblastosis virus.

Virus gene expression in rat cells transformed by either avian myelocytomatosis virus strain MC29 or avian erythroblastosis virus has been studied by biological and biochemical methods. In the clones examined, virus-specific sequences were found to be transcribed into RNA and, in most clones, the characteristic gag-related proteins could be identified. The transformed rat cells were fused to permissive chick cells and the rescued virus was shown to transform both chick embryo fibroblasts and the appropriate haemopoietic cell type in chick bone marrow cultures. These results clearly demonstrate that, as with the non-defective avian sarcoma viruses, the genetic information responsible for transformation by the defective avian leukaemia viruses can be expressed in non-permissive mammalian host cells as well as in permissive avian cells.

Expression of cellular homologues of retroviral onc genes in human hematopoietic cells.

Total cellular poly(A)-enriched RNA from a variety of fresh human leukemic blood cells and hematopoietic cell lines was analyzed for homology with molecularly cloned DNA probes containing the onc sequence of Abelson murine leukemia virus (Ab-MuLV), Harvey murine sarcoma virus (Ha-MuSV), simian sarcoma virus (SSV), and avian myelocytomatosis virus strain MC29. Results with the fresh blood cells paralleled those obtained with the cell lines. With Ab-MuLV and Ha-MuSV, multiple RNA bands were visualized in all cell types examined without significant variation in the relative intensities of the bands. When SSV was used as the probe, expression of related onc sequences was absent in all of the hematopoietic cell types examined except for one neoplastic T-cell line (HUT 102), which produces the human T-cell leukemia (lymphoma) retrovirus HTLV. In this cell line, a single band (4.2 kilobases) was observed. With MC29 as the probe, a single band of 2.7 kilobases was visualized in all cell types examined with only a 10 to 2-fold variation in intensity of hybridization. An exception was the promyelocytic cell line, HL60, which expressed approximately 10-fold more MC29-related onc sequences. With induction of differentiation of HL60 with either dimethyl sulfoxide or retinoic acid, a marked diminution in the amount of the MC29-related, but not the Ab-MuLV-related, onc message was observed.

Isolation and biochemical characterization of partially transformation-defective mutants of avian myelocytomatosis virus strain MC29: localization of the mutation to the myc domain of the 110,000-dalton gag-myc polyprotein.

Recently, we isolated three mutants of MC29 virus which, although able to transform fibroblasts with the same efficiency as wild-type MC29, were 100-fold less efficient at transforming macrophages. In this study we found that MC29-transformed quail producer cell line Q10 was able to generate these partially transformation defective mutants at a high frequency. Using tryptic peptide mapping, we determined that the smaller gag-myc polyproteins encoded by the transformation-defective viruses had lost myc-specific tryptic peptides. This suggested that the mutations which resulted in the transformation-defective viruses being inefficient at transforming macrophages were located in the v-myc sequence and thus directly implicated v-myc and the gag-myc polyprotein in transformation by MC29.

Mutants of avian myelocytomatosis virus with smaller gag gene-related proteins have an altered transforming ability.

Avian myelocytomatosis virus strain MC29 is a replication-defective avian oncovirus which in newborn chickens causes myelocytomatosis and liver and kidney tumours. In vitro infection of bone marrow cells gives rise to colonies of transformed macrophage-like cells, and cloned viruses is also capable of transforming fibroblasts. The genome of MC29 contains cellular sequences which are closely related to those in other defective leukaemia viruses with similar transforming spectra. Consequently, these cellular sequences have been postulated to represent a new oncogene which has been designated mac, for macrophage transformation. MC29-transformed cells contain a gag gene-related protein of a 110,000 molecular weight (MW) (p110), which by tryptic peptide analysis has been shown to be a fusion product comprised of a gag gene-derived sequences and sequences which are presumed to be coded by the adjacent mac gene. These findings suggest that this protein may be implicated in transformation by MC29. We now describe three mutants of MC29 and synthesize smaller gag gene-related proteins. These mutants have an altered ability to transform bone marrow cells but not fibroblasts. This demonstrates for the first time a direct involvement of the p110 protein of MC29 in transformation.

Transfection by DNAs of avian erythroblastosis virus and avian myelocytomatosis virus strain MC29.

Chicken embryo fibroblasts and NIH 3T3 mouse cells were transformable by DNAs of chicken cells infected with avian myelocytomatosis virus strain MC29 or with avian erythroblastosis virus. Transfection of chicken cells appeared to require replication of MC29 or avian erythroblastosis virus in the presence of a nontransforming helper virus. In contrast, NIH 3T3 cells transformed by MC29 or avian erythroblastosis virus DNA contained only replication-defective transforming virus genomes.

Transformation of mammalian cells by avian myelocytomatosis virus and avian erythroblastosis virus

Clones of rat cells transformed by avian myelocytomatosis virus strain MC29 and avian erythroblastosis virus have been isolated. These cells display an altered morphology and are able to form colonies in soft agar. Focus-forming virus can be rescued from the transformed rat cells by fusion with chick cells.

Transformation parameters in chicken fibroblasts transformed by AEV and MC29 avian leukemia viruses

Infection of chicken fibroblasts with avian erythroblastosis virus (AEV) strain ES4 or with avian myelocytomatosis virus strain MC29 leads to a rapid morphological transformation of most cells. AEV-transformed fibroblasts are similar to Rous sarcoma virus (RSV)-transformed fibroblasts in that they exhibit microvilli at their surface, show a disappearance of actin cables, are agglutinable by lectins, and show a decrease in LETS protein and an increase in the rate of hexose uptake. They also elicit slightly increased levels of cell-associated proteolytic activity, but show no increase in the fibrinolytic activity of the harvest fluids. In addition, as shown previously, they are capable of anchorage-independent growth and of sarcoma induction. In contrast, MC29-transformed fibroblasts express a different pattern of transformation parameters. They are similar to both RSV- and AEV-transformed fibroblasts in that they are morphologically transformed, show a disappearance of actin cables and are agglutinable by lectins. They also elicit surface alterations which consist of bleb-like protrusions rather than of microvilli, and are capable of anchorage-independent growth. They are strikingly different from RSV- and AEV-transformed cells, however, in that they express normal levels of LETS protein and elicit no increase in the rate of hexose uptake or in proteolytic activity. They are not sarcomagenic although they show an accelerated growth rate in culture. In conjunction with the finding that MC29 and AEV do not contain sequences related to the fibroblast-transforming src gene of RSV, these results raise the possibility that MC29 and perhaps also AEV transform fibroblasts by a mechanism different from RSV.

Avian myelocytomatosis and erythroblastosis viruses lack the transforming gene src of avian sarcoma viruses

Using labeled cDNA specific for the detection of the src gene of avian sarcoma viruses, we find that avian myelocytomatosis virus strain MC29 and avian erythroblastosis virus strain ES4 lack nucleotide sequences related to the src gene. Furthermore, chicken fibroblasts as well as hematopoietic cells, infected and transformed with these viruses, show no enhanced level of transcription of the cellular nucleotide sequences related to the src gene of avian sarcoma viruses. These two viruses may thus contain their own transforming gene(s) or induce cellular genes unrelated to the src-like cellular sequences.