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M A Baluda

Publications and source records attributed to M A Baluda.

At least 55 records · Page 3Linked to original sources
PubMed1980

Characterization of avian myeloblastosis-associated virus DNA intermediates.

The major species of unintegrated linear viral DNA identified in chicken embryonic fibroblasts infected with either the avian myeloblastosis-associated viruses (MAV-1, MAV-2) or the standard avian myeloblastosis virus complex (AMV-S) has a mass of 5.3 X 10(6) daltons. An additional minor DNA component observed only in AMV-S-infected cells has a mass of 4.9 X 10(6) daltons. The unintegrated linear viral DNAs and integrated proviruses of MAV-1 and MAV-2 have been analyzed by digestion with the restriction endonucleases EcoRI and HindIII. MAV-2 lacks a HindIII site present in MAV-1. These fragments have been compared to those generated by EcoRI and HindIII digestion of linear viral DNAs of AMV-S. Restriction enzyme digestion of AMV-S viral DNA produced unique fragments not found with either MAV-1 or MAV-2 viral DNAs. The major viral component present in AMV-S stocks has the HindIII restriction pattern of MAV-1. Restriction enzyme analysis of the 5.3 X 10(6)-dalton unintegrated MAV viral DNAs and their integrated proviruses suggests that the DNAs have a direct terminal redundancy of approximately 0.3 megadaltons and integrate colinearly with respect to the unintegrated linear DNA.

Avian Leukosis Virus↗
PubMed1980

Avian myeloblastosis virus proteins in leukemic chicken myeloblasts.

We have analyzed the avian myeloblastosis virus proteins in two types of leukemic myeloblasts: established myeloblastic cell lines (DU 1765 and DU 11157) and leukemic myeloblasts obtained from the peripheral blood of a leukemic C/E Spafas chicken (no. 21957). Using monospecific antisera for immunoprecipitation and polyacrylamide gel electrophoresis, we have detected gag gene-related proteins in the myeloblasts. The DU 1765 and DU 11157 cells contained a p100 protein which possessed antigenic determinants of the viral proteins p27, p19, p15, and p12. The p100 was not found in leukemic myeloblasts from Spafas chickens, and pulse-chase experiments showed that the p100 was not a precursor for the viral proteins. However, the p100 is present in uninfected line 15 chicken embryos. A pr76-like protein was identified in DU 1765 cells but migrated slightly further into gels than the pr76 of Spafas-derived leukemic myeloblasts. The Spafas-derived myeloblasts produced a pr60, whereas the DU 1765 cells contained instead a related protein of 62,000 daltons. Using anti-avian myeloblastosis virus gp85 sera, a glycoprotein of 120,000 daltons (gp120) was detected in all the tested leukemic myeloblasts. The gp120 was also present, in low amounts, in uninfected embyonic spleen and yolk sac cells. The anti-gp85 sera also precipitated a 27,000-dalton protein (h27) in these same cells. Both the gp120 and h27 could not be detected in either uninfected or myeloblastosis-associated virus-infected fibroblasts. Limited peptide hydrolysis revealed that h27 is different from the viral structural protein p27. In conclusion, monospecific antisera for gag and env gene products of avian myeloblastosis virus did not precipitate any unique or aberrant avian myeloblastosis virus protein from leukemic myeloblasts.

Animals↗
PubMed1980

DNA of avian myeloblastosis-associated virus type 2 integrates at multiple sites in the chicken genome.

The cellular sites of integration of the avian myeloblastosis-associated virus type 2 (MAV-2) DNA have been examined by Southern blot analysis of cellular DNA from infected cloned and uncloned chicken embryonic fibroblasts. Provirus-cell juncture fragments were not detected in restriction enzyme digests of DNA from MAV-2-infected uncloned cells. However, each MAV-2-infected cell clone examined produced a unique set of junctive bands. Thse findings indicate that multiple sites of integration exists for MAV-2 proviruses in cellular DNA.

Animals↗
PubMed1980

Identification of the avian myeloblastosis virus genome. I. Identification of restriction endonuclease fragments associated with acute myeloblastic leukemia.

The proviral DNA of chicken peripheral blood leukemic myeloblasts was analyzed by restriction endonuclease digestion and Southern blotting. Two restriction endonuclease-generated fragments, an EcoRI 2.2-megadalton (Md) and a HindIII 2.6-Md fragment, were present upon enzyme cleavage of all leukemic myeloblast DNA preparations in addition to endogenous or helper-specific fragments. Neither of these fragments was derived from viral DNA of the two known myeloblastosis-associated viruses (MAV-1 and MAV-2). In contrast, DNA similarly treated from the erythrocytes of leukemic chickens showed only small amounts of the two avian myeloblastosis virus-specific fragments, whereas the helper virus-specific fragments were present in the amount seen in MAV-producing chicken embryo fibroblasts. The appearance of the EcoRI 2.2-Md and HindIII 2.6-Md specific fragments in all leukemic myeloblast DNA preparations indicates they are presumably part of the leukemogenic genome that must be present to induce acute myeloblastic leukemia.

Animals↗
PubMed1980

Identification of the avian myeloblastosis virus genome. II. Restriction endonuclease analysis of DNA from lambda proviral recombinants and leukemic myeoblast clones.

Two lambda proviral DNA recombinants were characterized with a number of restriction endonucleases. One recombinant contained a complete presumptive avian myeloblastosis virus (AMV) provirus flanked by cellular sequences on either side, and the second recombinant contained 85% of a myeloblastosis-associated virus type 1 (MAV-1)-like provirus with cellular sequences adjacent to the 5' end of the provirus. Comparing the restriction maps for the proviral DNAs contained in each lambda hybrid showed that the putative AMV and MAV-1-like genomes shared identical enzyme sites for 3.6 megadaltons beginning at the 5' termini of the proviruses with respect to viral RNA. Two enzyme sites near the 3'-end of the MAV-1-like provirus were not present in the putative AMV genome. We also examined a number of leukemic myeloblast clones for proviral content and cell-provirus integration sites. The presumptive AMV provirus was present in all the leukemic myeloblast clones regardless of the endogenous proviral content of the target cells or the AMV pseudotype used for conversion. Multiple cellular sites were suitable for integration of the putative AMV genome and the helper genomes. The proviral genomes were all integrated colinearly with respect to linear viral DNA.

Animals↗
PubMed1979

Integration of Rous-associated virus type O provirus in susceptible chicken cells.

The number of viral genome equivalents per haploid cell genome was determined in normal chicken embryos from three selected chicken lines and in cultured fibroblasts (CEF) from these embryos. The cellular concentration of endogenous proviral DNA is similar in embryos from chickens of lines SPAFAS, 7, 15, 7 x 15, and 100. The concentration of proviral DNA is not affected by in vitro cultivation in CEF from lines that do not spontaneously produce virus, nor in CEF from line 7, which lacks receptors for Rous-associated virus type 0 (RAV-0). There is, however, a restricted increase in the number of integrated proviral genome equivalents in CEF from line 7 x 15, which produces RAV-0 and can support replication of this virus, and in CEF from line 15 experimentally infected with RAV-0.

Animals↗
PubMed1977

Incomplete viral genome in a non-virogenic mouse tumour cell line (RVP3) transformed by Prague strain of avian sarcoma virus.

Two cell lines, RVP3 and RVA4, derived originally from mouse tumors induced by the Prague and Schmidt-Ruppin strain of RSV, respectively, were studied. tall attempts failed to induce infectious virus production in RVP3 cells by fusion with chicken fibroblasts even if the cells were infected with avian leukosis viruses. Also, attempts to rescue the viral genome by transfection were unsuccessful. RVP3 cells harboured 31-45% of the viral genome sequences, as was shown by molecular hybridization, and therefore they were designated cryptovirogenic. The tumour cell line RVA4 did not contain any detectable viral sequences. The significance of the detection of the incomplete Rous virus genome sequences in mammalian cells is discussed.

Animals↗
PubMed1977

Variations in integration site of avian oncornaviruses in different hosts.

We examined the integration site of avian oncornaviruses in the genome of different hosts with respect to the repetitive frequency of the cellular DNA sequences adjacent to the integrated proviral DNA. The following systems were studied: avian sarcoma virus (B-77) and avian leukosis virus (Rous-associated virus-61) in cultured duck embryonic cells and B-77 in cultured mouse 3T3 cells. These systems represent different host responses to viral infection, i.e., one in which both cellular transformation and viral replication occur (B-77-infected duck cells), one in which viral replication, but not transformation, occurs (Rous-associated virus-61-infected duck cells), and one in which transformation, but not viral replication, occurs (B-77-infected 3T3 cells). Two sequential hybridizations were used. First, large denatured DNA fragments (2.8 X 10(6) daltons) were reassociated to different C0t (mole-seconds per liter) values. Next, DNA remaining single stranded at different C0t values was isolated by hydroxylapatite column chromatography, immobilized on nitrocellulose filters, and hybridized with an excess of 3H-labeled 35S viral RNA to titrate the concentration of proviral DNA. Results show that B-77 sarcoma virus and Rous-associated virus-61 integrate in the unique region of duck DNA, whereas B-77 proviral DNA is associated with both repeated and unique host DNA sequences in transformed mouse 3T3 cells.

Animals↗
PubMed1977

Homogeneity and complexity of avian oncornavirus proviral DNA determined by molecular hybridization.

The homogeneity of DNA complementary to the 35S RNA subunit of avian myeloblastosis virus (AMV) has been demonstrated by single or multistep hybridization. For multistep hybridizations, 35S AMV RNA was preselected for its ability to hybridize either to unfractionated leukemic DNA or to leukemic DNA enriched for unique or for reiterated sequences. These experiments indicate that the viral genome is complementary to DNA sequences with a low reiteration frequency. Competition experiments confirm the absence of fast-hybridizing sequences in viral DNA. Computer analyses of the data reveal that there are two to four copies of viral DNA in infected cells.

Animals↗
PubMed1977

Integration of proviral DNA in chicken cells infected with Schmidt-Ruppin Rous sarcoma virus is not enhanced by DNA repair.

The effect DNA repair might have on the integration of exogenous proviral DNA into host cell DNA was investigated by comparing the efficiency of proviral DNA integration in normal chicken embryonic fibroblasts and in chicken embryonic fibroblasts treated with UV or 4-nitroquinoline-1-oxide. The cells were treated with UV or 4-nitroquinoline-1-oxide at various time intervals ranging from 6 h before to 24 h after infection with Schmidt-Ruppin strain A of Rous sarcoma virus. The chicken embryonic fibroblasts were subsequently cultured for 18 to 21 days to ensure maximal integration and elimination of nonintegrated exogenous proviral DNA before DNA was extracted. Integration of proviral DNA into the cellular genome was quantitated by hybridization of denatured cellular DNA on filters with an excess of (3)H-labeled 35S viral RNA. The copy number of the integrated proviruses in normal cells and in infected cells was also determined from the kinetics of liquid RNA-DNA hybridization in DNA excess. Both RNA excess and DNA excess methods of hybridization indicate that two to three copies of the endogenous provirus appear to be present per haploid normal chicken cell genome and that two to three copies of the provirus of Schmidt-Ruppin strain A of Rous sarcoma virus become integrated per haploid cell genome after infection. The copy number of viral genome equivalents integrated per cell treated with UV or 4-nitroquinoline-1-oxide at different time intervals before or after infection did not differ from the copy number in untreated but infected cells. This finding supports our previous report that the integration of oncornavirus proviral DNA is restricted to specific sites in the host cell DNA and suggests a specific mechanism for integration.

4-Nitroquinoline-1-oxide↗
PubMed1976

Restricted addition of proviral DNA in target tissues of chickens infected with avian myeloblastosis virus.

Proviral DNA is synthesized within an hour after infection of chicken cells with an avian oncornavirus and is integrated into nuclear cellular DNA within a short time. The viral DNA appears to be synthesized as double-stranded molecules of approximately 6 X 10(6) daltons some of which are converted into supercoiled cricles perhaps as a requisite for integration. The endogenous v-DNA in normal chicken cells and both the endogenous and amv v-DNA in leukemic chicken myeloblasts are covalently linked with chromosomal DNA. There is no detectable free DNA either circular or linear present in leukemic cells several weeks after infection. The endogenous v-DNA which is transmitted vertically from parents to offspring is uniformly and stably distributed in all chicken organs. There are about 1-2 copies of endogenous provirus per haploid genome of all normal cells. This DNA is very closely related to RAV-O RNA. After infection with AMV it seems that target cells such as leukemic myeloblasts, RBC and nephroblasts acquire complete copies of AMV DNA. Interestingly, only these target cells can be converted to neoplastic cells in the chicken as well as in vitro. The target cells acquire 1-2 copies of AMV specific DNA per haploid genome in addition to the endogenous v-DNA. All the available evidence shows that leukemic and kidney tumor cells have acquired AMV v-DNA. It remains to be elucidated whether the newly added viral DNA is alone responsible for neoplastic changes or does so in conjunction with endogenous viral information.

Animals↗
PubMed1976

Evidence for tandem integration of avian myeloblastosis virus DNA with endogenous provirus in leukemic chicken cells.

The integration site of avian myeloblastosis virus (AMV) proviral DNA in DNA from leukemia chicken myeloblasts has been studied by three sequential nucleic acid hybridizations that can localize the proviral DNA according to the repetitiveness of the adjacent cellular DNA regions. First, large denatured cellular DNA fragments (2.1 x 10(6) daltons) were reassociated and fractionated according to sequence reiteration frequenct. Next, DNA remaining single-stranded in each fraction was immobilized on nitrocellulose filters hybridized with an excess of unlabeled 70S RNA from Rous-associated virus-0 to saturate the endogenous proviral DNA sequences.

Animals↗
PubMed1975

Host induced alteration of avian sarcoma virus B-77 genome.

The genome of an avian oncornavirus was altered after infection of a heterologous host. This was studied with avian sarcoma virus B-77 in duck embryonic fibroblasts (DEF) and chicken embryonic fibroblasts (CEF). To detect alteration of the viral genome, we hybridized 35S B-77 RNA with normal duck DNA by either one of two techniques:when viral RNA was in excess and when DNA was in excess. The RNA of B-77 passaged only in gs minus chf minus CEF does not have homology with duck DNA. However, after several passages of B-77 through DEF the viral genome acquired duck specific RNA sequences. After 4 and 10 passages, B-77 RNA acquired 2.2 and 6.6%, respectively, complementarity to normal duck DNA. The duck specific RNA sequences were found to be covalently linked to the B-77 RNA genome. Also, the host specific sequences acquired by the virus appear to be from a region of the duck DNA which is repeated four to six times per cell. After 5 back passages in CEF some of the duck specific RNA sequences in the viral genome were lost.

Animals↗
PubMed1975

Studies on characterization of the integration sites of avian RNA tumor virus-specific DNA.

A sequential hybridization procedure is described which allows the integration sites of viral-specific DNA to be characterized according to their reassociation kinetics. In addition, this approach enables us to estimate the size of the integrated viral DNA. Endogenous virus sequences in normal cells appear to be associated with cell sequences reiterated 1200 times, and each integration unit is approximately equal to one 35S RNA subunit. In AMV-infected cells, the additional AMV-specific DNA sequences reassociate as if they were integrated adjacent to unique cellular DNA or in tandem with the endogenous viral DNA sequences.

Animals↗
PubMed1975

RNA of simian sarcoma-associated virus type 1 produced in human tumor cells.

Simian sarcoma-associated virus type 1 propagated in human rhabdomyosarcoma cells exhibited characteristics typical of oncornaviruses but seemed to have several aberrant properties. It had a buoyant density of 1.14 g/cm3, had RNA-dependent DNA polymerase activity, seemed to be labile to high salt concentrations, and contained little 50 to 60S RNA but relatively large amounts of human ribosomal RNA. In addition to 50 to 60S RNA, purified virions contained smaller RNA molecules with sedimentation coefficients of 28 to 30S, 18 TO 20S, and 4 to 10S. Unlike the 50 to 60S RNA species, the smaller virion-associated RNAs lacked polyadenylic acid, and the 28 to 30S RNA had an average base composition similar to that of human ribosomal RNA. Upon heat denaturation, the native 50 to 60S RNA genome yielded polyadenylic acid-containing 28 to 30S subunits that degraded in to 18 to 20S molecules upon further heat treatment. The 50 to 60S viral RNA had a guanine plus cytosine content of 56%.

Animals↗
PubMed1975

Interspersion of sequences in avian myeloblastosis virus rna that rapidly hybridize with leukemic chicken cell DNA.

Liquid hybridization of progressively smaller fragments (35S, 27S, 15.5S, 12.5S, and 8S) of poly(A)-selected avian myeloblastosis virus RNA with excess DNA from leukemic chicken myeloblasts revealed that all sizes of RNA contained sequences complementary to both slowly and rapidly hybridizing cellular DNA sequences. Apparently, the RNA sequences which hybridize rapidly with excesses of cellular DNA are not restricted to any one region of the avian myeloblastosis virus 35S RNA. Instead, they appear to be randomly distributed over the entire 35S avian myeloblastosis virus RNA molecule with some positioned within 200 nucleotides of the poly(A) tract at the 3' end of the RNA.

Animals↗