Search PubMed⌕ Search

Biomedical subjects

S Cory

Publications and source records attributed to S Cory.

At least 91 records · Page 5Linked to original sources
PubMed1988

Comparison of expression in hemopoietic cells by retroviral vectors carrying two genes.

In order to identify factors that influence expression by retroviral vectors in hemopoietic cells, we have compared viral RNA levels in cells infected with several different recombinant viruses. All of the vectors tested carry the neomycin resistance gene and provide for the insertion of a second gene which, in these studies, comprised sequences from the myc or myb oncogenes or the gene encoding granulocyte-macrophage colony-stimulating factor. The vectors utilize two different strategies for the coexpression of the two genes: alternate splicing and the use of a separate internal promoter. We found that expression in hemopoietic cells could be increased by substituting sequences from the myeloproliferative sarcoma virus long terminal repeat for those of the Moloney murine leukemia virus long terminal repeat. However, none of the vectors examined was able to express a second gene at levels equivalent to those achieved by the parental vectors carrying only the neomycin resistance gene. The reasons for this varied with the different vectors and included inefficient splicing and/or a reduction in the level of unspliced transcripts upon insertion of a second gene. Although the basis of the latter phenomenon is not clear, it is probably related to the position--near the 5' long terminal repeat--at which the second gene was inserted, since insertion of the same genes near the 3' end of another vector had no effect on viral RNA levels. In an attempt to circumvent some of these problems, we constructed a vector that employs an internal beta-actin promoter. Although this vector could express granulocyte-macrophage colony-stimulating factor sequences in a responsive hemopoietic cell line, the level of granulocyte-macrophage colony-stimulating factor produced was disappointingly low. The results from these studies suggest approaches to the design of improved vectors for effective expression of genes in hemopoietic cells.

Animals↗
PubMed1988

Transformation of bone marrow cells from E mu-myc transgenic mice by Abelson murine leukemia virus and Harvey murine sarcoma virus.

Transgenic mice harboring a c-myc gene subjugated to the immunoglobulin heavy chain enhancer offer a unique opportunity to investigate whether deregulated myc expression potentiates the transformation of B lymphoid cells by other oncogenes. By assessing colony formation in semi-solid medium, we have compared the potential of bone marrow cells from E mu-myc mice and their normal littermates for transformation by Harvey murine sarcoma virus and Abelson murine leukemia virus. E mu-myc bone marrow yielded more lymphoid colonies than normal marrow after infection with Harvey virus. The increased transformation frequency may reflect increased clonogenicity due to complementation between myc and ras and/or the increased number of pre-B cells in E mu-myc marrow. Surprisingly, however, the number of lymphoid colonies induced by Abelson virus was not enhanced. Our interpretation of these results is that the primary Abelson target is more primitive than the pre-B cells expressing the E mu-myc transgene and is therefore not present at increased frequency in the E mu-myc marrow. The cells from most virus-infected E mu-myc colonies failed to grow indefinitely when placed in liquid culture in the absence of a feeder layer. Thus expression of a deregulated c-myc gene together with either v-Ha-ras or v-abl does not ensure fully autonomous growth of early B lymphoid cells.

Abelson murine leukemia virus↗
PubMed1988

Growth of E mu-myc transgenic B-lymphoid cells in vitro and their evolution toward autonomy.

Constitutive expression of the c-myc oncogene in the B lymphoid cells of E mu-myc transgenic mice promotes cellular proliferation and predisposes to lymphomagenesis. To delineate further how the pre-B cells of prelymphomatous mice have been altered, we have investigated their growth in vitro. They were not autonomous, since they died rapidly when cultured without feeder cells. When cultured on bone marrow stromal cells, E mu-myc cells initially grew to only slightly higher densities than B lineage cells from normal mice, but were larger and showed more cell cycle activity. After 14 weeks, all cultures appeared oligo- or monoclonal, as judged by analysis of Ig gene rearrangements. While the growth of the normal cells was constant, the E mu-myc cells started to grow to 10-fold higher densities after 14-20 weeks, implying a reduced requirement for growth factors. They remained dependent on feeder cells and were non-tumorigenic. By 25 weeks, however, the one remaining culture had become feeder-independent and tumorigenic. We suggest that deregulated c-myc expression in B cell precursors increases the rate of cell turnover, thereby increasing the frequency of genetic changes conductive to autonomous growth.

Animals↗
PubMed1988

bcr-abl oncogene renders myeloid cell line factor independent: potential autocrine mechanism in chronic myeloid leukemia.

In chronic myeloid leukemia (CML), a chromosome translocation has fused the bcr gene to the c-abl oncogene, such that a chimeric bcr-abl polypeptide can be made. To explore the biological properties of bcr-abl and compare them with those of the Abelson virus (AMuLV) transforming gene (gag-v-abl), we have used either a synthetic bcr-v-abl gene that mimics the translocation product or, in some experiments, a bcr-c-abl cDNA. A new retroviral vector was used to introduce the genes into the factor-dependent myeloid line FDC-P1. Both bcr-abl and v-abl efficiently rendered the myeloid cells factor independent and tumorigenic. Their fully autonomous growth may be due to the myeloid growth factor interleukin-3 (IL-3) made in small amounts by the infected cells. Hence autocrine factor production may feature in CML development and Abelson virus transformation.

Animals↗
PubMed1987

Generation of altered transcripts by retroviral insertion within the c-myb gene in two murine monocytic leukemias.

Two murine monocytic leukemia cell lines, WEHI-265 and WEHI-274, were found to carry a rearranged c-myb gene. The rearrangements are due to insertion of a deleted Moloney murine leukemia virus (Mo-MLV) provirus in the 5' region of the c-myb gene and thus are similar to rearrangements in the ABPL tumors (G. L. C. Shen-Ong, M. Potter, J. F. Mushinski, S. Lavu, and E. P. Reddy, Science 226:1077-1080, 1984). In each cell line, the retroviral insertion has induced high levels of two aberrant RNA species, which, as in the ABPL tumors (G. L. C. Shen-Ong, H. C. Morse, M. Potter, and J. F. Mushinski, Mol. Cell. Biol. 6:380-392, 1986), contain both viral (Mo-MLV) and cellular (myb) sequences. Both species lack the sequences encoding the amino terminus of the c-myb protein and thus could encode a protein which, like the v-myb gene products (and the predicted ABPL myb proteins), is truncated at the amino terminus. We have found that the larger (5.3 kilobase [kb]) and more abundant of the tumor-specific myb RNAs was predominantly nuclear, while the smaller species (3.9 kb) was cytoplasmic. Furthermore, our data imply that the 3.9-kb RNA was derived from the 5.3-kb RNA by an additional splice which utilized a cryptic splice acceptor site within the viral gag sequences. On the basis of subcellular distribution and predicted translational potential, we conclude that the 3.9-kb RNA is probably the mRNA which encodes a truncated myb protein. We also show that, due to different insertion points in W265 and W274, the W274 myb RNAs contained sequences from a c-myb exon upstream of the exons represented in the W265 (and ABPL) RNAs. The significance of our findings with regard to transformation by myb in these tumors is discussed.

Base Sequence↗
PubMed1987

Murine c-myc retroviruses alter the growth requirements of myeloid cell lines.

Recombinant retroviruses encoding a murine c-myc gene were utilised to explore the consequences of constitutive c-myc expression for interleukin 3-dependent murine myeloid cell lines. The c-myc virus-infected cells exhibited a diminished requirement for growth factor and serum when grown in soft agar, but no factor-independent cells could be isolated and the cells were not tumorigenic. Thus deregulated expression of a c-myc gene at physiological levels apparently renders myeloid cells more responsive to growth factors but cannot abrogate this requirement or render the cells malignant.

Animals↗
PubMed1987

Expression of genes transferred to haemopoietic stem cells by recombinant retroviruses.

We have compared several recombinant retroviruses carrying the selective marker NeoR for gene delivery to murine haemopoietic stem cells. Provirus content and expression was assessed in animals reconstituted with infected bone marrow cells. With viruses bearing only a NeoR gene, viral RNA was readily detectable both in spleen colonies and in the haemopoietic tissues of long-term reconstituted mice. Thus, the progeny of infected primitive multipotential cells are competent to express integrated proviruses. The level of expression was significantly greater for proviruses carrying a modified long terminal repeat in which the Moloney enhancer had been replaced with that from the myeloproliferative sarcoma virus. With viruses bearing the c-myc gene in addition to NeoR, however, no viral RNA could be detected in spleen colonies harbouring proviral inserts, even though these viruses are expressed in vitro within haemopoietic cells. The implications of these results for gene transfer to haemopoietic stem cells are discussed.

Animals↗
PubMed1986

The c-myc oncogene perturbs B lymphocyte development in E-mu-myc transgenic mice.

Transgenic mice bearing a c-myc oncogene subjugated to the lymphoid-specific immunoglobulin heavy chain enhancer (E mu) develop clonal B lymphoid malignancies, but most young E mu-myc mice lack malignant clones. Their prelymphomatous state has allowed us to examine how constitutive c-myc expression influences B cell development. We find that early stages are overrepresented, even before birth. Pre-B cells of polyclonal origin increase greatly, while B cells develop in reduced number. Both the pre-B and the B cells appear to be in an active state, since they are larger than normal and a greater fraction are in the cell cycle. Enforced myc expression has thus favored proliferation over maturation. Hence, a normal function of c-myc may be to regulate differentiation as well as to promote cell cycling.

Animals↗
PubMed1985

Myc oncogene activation in B and T lymphoid tumours.

The chromosome translocations characteristic of certain B lymphoid tumours associate the myc oncogene and immunoglobulin loci. The typical t(12;15) in murine plasmacytomas and analogous t(14;8) in Burkitt lymphomas couple the myc coding region to one of the switch recombination regions within the immunoglobulin heavy (H) chain locus; hence the switch machinery may promote some translocations. Significantly, translocation induces constitutive myc expression, the untranslocated myc allele remaining silent. The predilection for breakpoints near the 5' end of the c-myc gene may reflect selection for altered myc regulation. In most tumours, the stimulatory effect of the H locus context is not understood, but an H locus enhancer participates in some tumours, including one displaying a novel transposition. The variant (6;15) translocations found in about 15% of plasmacytomas involve the myc band and the region of chromosome 6 where the kappa locus lies. The t(6;15) is shown here to represent an exchange between C kappa and a chromosome 15 locus (designated pvt-1) which lies unexpectedly far from c-myc. The association of myc expression with pvt-1 alterations suggest that myc can be activated at a distance. Myc has also been implicated in some T lymphomas by detection of proviral inserts near myc and also, surprisingly, within the pvt-1 locus. Inserts near myc appear to activate its expression via the retroviral enhancer.

Animals↗
PubMed1985

Variant (6;15) translocations in murine plasmacytomas involve a chromosome 15 locus at least 72 kb from the c-myc oncogene.

The variant (6;15) translocations in murine plasmacytomas join the myc oncogene-bearing band of chromosome 15 and the immunoglobulin kappa band of chromosome 6. We recently cloned a region from chromosome 15 linked to C kappa and have now used probes from that region to define the major locus of plasmacytoma variant translocations, which we denote pvt-1. In five of nine plasmacytomas we analysed, the 6;15 translocation resulted from reciprocal recombination between the C kappa locus and a 4.5-kb region of pvt-1. Moreover, nearby we located the region shown by others to have undergone a complex (15;12;6) translocation in plasmacytoma PC7183. All the chromosome 6 breakpoints fell between 1 and 3 kb 5' to C kappa but only two were near J kappa genes. Thus the J kappa -C kappa region appears to be a recombination 'hot spot' in lymphocytes, but the breaks are unlikely to be mediated via V/J recombination enzymes. Comparison of a cloned 108-kb region across pvt-1 and another of 52 kb across c-myc established that the pvt-1 breakpoints lie at least 72 kb from the c-myc promoters. Since c-myc is expressed at a substantial level, the 6;15 translocation apparently activates c-myc. Activation may occur directly, at a remarkable distance along the chromosome, or indirectly, via a putative pvt-1 gene product.

Animals↗
PubMed1985

Transposition of the immunoglobulin heavy chain enhancer to the myc oncogene in a murine plasmacytoma.

A novel mechanism of oncogene activation by transposition of a tissue-specific cellular enhancer is described. A rearranged c-myc oncogene was cloned from murine plasmacytoma ABPC17 in the expectation that it would reflect the t(6;15) chromosome translocation carried by this tumor. The rearrangement instead reflects an insertion 361 bp 5' to the c-myc gene on chromosome 15. The insert conveys a 2.3 kb segment of the immunoglobulin heavy (H) chain locus from chromosome 12. Since the insertion introduces the lymphoid-specific enhancer from the JH-Smu region and also disrupts a region implicated in normal c-myc control, it may account for the c-myc transcription observed in ABPC17. The structure of the transposed segment and a corresponding deletion in the JH-Smu region suggests that the transposition reflects a complex recombination between chromosomes 15 and 12. Since the t(6;15) breakpoint is not near c-myc, chromosome 15 must have undergone an independent exchange with chromosome 6.

Alleles↗
PubMed1984

Translocation of the myc cellular oncogene to the immunoglobulin heavy chain locus in murine plasmacytomas is an imprecise reciprocal exchange.

The 15;12 translocations in murine plasmacytomas represent recombination of the myc oncogene with the immunoglobulin CH locus, often within CH switch recombination (SH) regions. Chromosome junctions cloned from four plasmacytomas confirmed that the translocation generates reciprocal CHSH-myc and 5'myc-SH structures. H locus targets included S alpha fused to Smu, Smu fused to S gamma 2b, and a germline S alpha region. The nature of two H locus targets suggests that the target need not be highly active transcriptionally. Switch recombination machinery is implicated in the translocation by the SH targets and by homology of certain c-myc breakpoints with normal switch recombination sites. Fusion regions revealed deletions, extraneous nucleotides, and one duplication. These results prompt a translocation model in which staggered single-stranded breaks on each chromosome are followed by single-strand excision or polymerization prior to ligation to the other chromosome.

Alleles↗
PubMed1984

Murine T lymphomas in which the cellular myc oncogene has been activated by retroviral insertion.

The myc oncogene is implicated here in T lymphocyte neoplasia. Cloning revealed a retroviral insert 0.7-1.3 kb 5' to c-myc in two T lymphomas induced by Soule murine leukemia virus and in a spontaneous T lymphoma ( Tikaut ) of an AKR mouse, a strain in which leukemogenesis involves recombinant retroviruses (MCF viruses). The tumor c-myc mRNAs appear normal but their level is approximately 5-fold higher than in most T lymphomas lacking c-myc rearrangement. Since each insert would be transcribed away from c-myc, its activation cannot involve the promoter of the long terminal repeat (LTR) but could reflect an enhancer, like that demonstrated within the Soule LTR. The Tikaut provirus has an MCF-like recombinant env gene and LTR sequence. MCF-like inserts were found near c-myc in seven of 31 other AKR T lymphomas; two lie 3' to c-myc and the five upstream are oriented away from c-myc. We conclude that a quarter of retrovirus-induced T lymphomas involve activation of c-myc, probably via the LTR enhancer.

Animals↗