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Endogenous New World primate retrovirus: interspecies antigenic determinants shared with the major structural protein of type-D RNA viruses of Old World monkeys.

A reverse transcriptase-containing virus has recently been isolated from a squirrel monkey (Saimiri sciureus). Molecular hybridization studies demonstrate that the squirrel monkey retrovirus (SMRV) is endogenous to this New World primate, yet lacks detectable nucleotide sequence homology with cellular DNAs of representative Old World primates or with the genomes of previously isolated Old World primate retroviruses. The 35,000-dalton major structural protein (p35) of SMRV was purified and shown to possess antigenic determinants distinct from those of known retroviruses. While SMRV was found to lack antigenic determinants broadly shared among mammalian type-C viruses, immunologic crossreactivity was demonstrated between SMRV p35 and the major structural protein (p26) of Mason-Pfizer monkey virus, a prototype type-D retrovirus of Old World monkeys. These findings support the concept that SMRV and Mason-Pfizer monkey virus are evolutionarily related, and raise the possibility that a progenitor of type-D retroviruses became genetically associated with primates at a very early time in their evolution.

Animals↗

Type D primate retroviruses: a review.

The prototype virus of the type D retroviruses is the Mason-Pfizer monkey virus (MPMV). MPMV was originally isolated from a breast carcinoma of a female rhesus monkey (an Old World monkey). MPMV is of obvious importance in that it is the only retrovirus thus far isolated from a mammary tumor of a primate and has been shown to have transforming potential for primate cells in vitro. Subsequent to the isolation of MPMV viruses morphologically and immunologically indistinguishable from MPMV have been isolated from normal placenta and lactating mammary glands of other rhesus monkeys in captivity. Recently, viruses morphologically resembling MPMV have been isolated from a langur monkey (another Old World monkey) and from squirrel monkeys (a New World monkey). Based on nucleic acid hybridization studies, the latter 2 viruses represent endogenous viruses in their species of origin, whereas MPMV appears to be a horizontally related to the langur monkey isolate. Studies on the immunological relatedness of the type D retroviruses have demonstrated interspecies cross-reactivities between the major internal and external proteins of the viruses. Furthermore, these viruses also share cross-reactivity of their major external glycoproteins with those of the type C baboon endogenous virus. These interspecies reactivities can also be demonstrated in natural sera from both imported and laboratory-bred monkeys. The demonstration of these interspecies cross-reactivities shared by distantly related primate retroviruses provides a means for detecting determinants that are representative of all primate retroviruses presently known and yet to be isolated and may provide new assays for detection of a human retrovirus.

Animals↗

Detection of simian type-C particles in Mason-Pfizer virus infected cultures.

A simian type-C virus has been detected in cultures chronically infected with Mason-Pfizer monkey virus (M-PMV). Simultaneous budding of M-PMV and type-C virus particles from the same cells was observed in cultures incubated at 37 or 40 degrees C. However, the frequency of such cells was greater in cultures grown at 40 degrees C. Although clusters of type-C viral buds were seen at the surface of the cells, extracellular mature type-C particles in cell pellets or concentrated virus preparations were very rarely found. The increase in frequency of type-C buds was found to be transitory since cultures adapted to growing at the high temperature demonstrated budding type-C particles only occasionally. Cultures producing type-C buds were found to contain, in addition to M-PMV antigens, serological activity with polyvalent antisera produced against multiple structural components of endogenous baboon virus (BV) or simian sarcoma virus (SiSV). The reactivity, however, was found not to be serologically related to the major SiSV P28 core protein.

Animals↗

Detection, quantitation, and characterization of the major internal virion antigen of the bovine leukemia virus by radioimmunoassay.

The major internal polypeptide of the bovine leukemia virus (BLV) was purified to homogeneity with the use of gel filtration and affinity chromatography. Like previous results, the protein had a molecular weight of 25,000 daltons as determined by electrophoresis in polyacrylamide gels with sodium dodecyl sulfate. More than 90% of the 125I-labeled protein was precipitated by bovine sera that reacted in immunofluorescence tests with acetone-fixed BLV-infected cells. In contrast, minimal precipitation (less than 5%) was observed with sera from 36 cattle in leukemia-free herds; these sera, negative by immunofluorescence, included six samples that had high titers of antibodies to the foamy-like bovine syncytia virus (BSV). Antisera prepared against several other oncornaviruses or the Mason-Pfizer monkey virus (M-PMV) did not bind the BLV p25 protein. Conversely, the labeled p30 polypeptides of several oncornaviruses tested did not react with bovine sera that had high titers of antibodies to BLV p25. Competitive radioimmunoassay(s) (RIA) also failed to detect cross-reactions between BLV p25 protein and the internal polypeptides of other mammalian and avian oncornaviruses, M-PMV, or foamy-like BSV. The RIA for BLV p25 antigen was also highly sensitive and specific for the detection and quantitation of the antigen in virus preparations and cell homogenates.

Animals↗

Expression of Mason-Pfizer and simian type C viruses in the presence of 5-iododeoxyuridine and dexamethasone.

Production of infectious Mason-Pfizer monkey virus (M-PMV) was enhanced after treatment of the CMMT cell line with 2.5 x 10(-5) M dexamethasone phosphate (DXM). The reverse transcriptase (RT) activity and infectivity titers of treated culture fluids were enhanced by five- and tenfold, respectively. Along with stimulation of M-PMV synthesis, a simian type C virus (SCV) was also detected by electron microscopic and RT analyses. The SCV was serologically related to the endogenous baboon type C virus. 5-iododeoxyuridine (IUDR) also activated the SCV in the CMMT cell line while significantly inhibiting the production of infectious M-PMV. The activation of endogenous SCV by IUDR or DXM was transitory, since removal of these compounds from the growth medium resulted in the disappearance of SCV buds and the related RT activity; however, low levels of specific viral structural proteins continued to be synthesized intracellularly. Similarly, the enhancement of M-PMV production seen with DXM was lost when the treated cells were subcultured for 2 weeks in the absence of the hormone.

Cell Line↗

Equine infectious anemia virus: evidence favoring classification as a retravirus.

Equine infectious anemia virus (EIAV) has a density of 1.154 g/cm3 in sucrose a high-molecular-weight RNA similar in size to Rauscher murine leukemia virus, and an internal virion reverse transcriptase that utilizes the synthetic RNA template poly(rA) but not the synthetic DNA template poly(dA), both with (dT)12 as primer. Although capable of utilizing manganese at low concentrations (approximately 0.1 mM), EIAV reverse transcriptase showed highest activity in the presence of 9 mM magnesium. The major protein of EIAV has a slightly lower molecular weight than the comparable protein of type C viruses and co-electrophoresed with 125I-labeled p25 of Mason-Pfizer monkey virus. A reference horse serum with antibodies to the major EIAV protein reacted only with EIAV and not with other type C or non-type C retraviruses. Reciprocally, a broadly reactive serum to type C virus p30s and specific sera to a variety of non-type C retraviruses did not react with EIAV. We recommend the inclusion of EIAV in the family Retraviridae.

Antigens, Viral↗

Apparent allosterism by avian myeloblastosis virus reverse transcriptase and E. coli DNA polymerase I.

A recent report (1) presented evidence for allosterism in reverse transcription by Mason-Pfizer monkey virus reverse transcriptase and by E. coli DNA polymerase I. Our experiments also demonstrate these apparent cooperative effects when synthesis is catalyzed by either avian myeloblastosis virus DNA polymerase, feline sarcoma virus DNA polymerase, or E. coli DNA polymerase I (large fragment). We show that the apparent cooperativity depends on the use of oligo(dT)12-18 as primer. However, if the polymerase reaction products are isolated chromatographically, then the polymerases obey classical Michaelis-Menten kinetics with respect to substrate and enzyme concentrations. These results suggest that the cooperative effects are an acid precipitation artifact. The results are also consistent with the enzyme operating by a distributive mechanism with the oligo(dT)12-18 primer.

Allosteric Regulation↗

Serological definition of the lentivirus group of retroviruses.

The major polypeptides of visna viruses and other lentiviruses have been isolated and shown to be closely related if not identical in radioimmunoassays. By this criterion the lentiviruses form a distinct group of retroviruses unrelated to spuma viruses, mammalian and avian retroviruses that cause tumors, and unclassified retroviruses of cattle and horses. Two sera obtained from goats immunized with Mason-Pfizer monkey virus or squirrel monkey virus reacted with visna p30. Additional data suggest that this reaction represents infection of goats with a lentivirus or a new retrovirus closely related to the lentiviruses.

Animals↗

Primate retroviruses: immunological cross-reactivity between major structural proteins of new and old world primate virus isolates.

The major 35,000-molecular-weight internal antigen (p35) of the squirrel monkey retrovirus (SMRV) was isolated and partially characterized. Immunological analysis of SMRV p35 led to the demonstration of antigenic determinants common to SMRV and the Mason-Pfizer monkey virus (MPMV). A broadly reactive competition immunoassay was developed utilizing antiserum to MPMV to precipitate 125I-labeled SMRV p35. Although the major structural proteins of MPMV and SMRV competed with equal efficiency in this assay, type B and type C oncornavirus proteins lacked detectable reactivity. Antibodies reactive with the major structural proteins of both MPMV and SMRV were observed in sera of several normal rhesus monkeys with known prior exposure to MPMV-infected animals. These findings demonstrate the ability of sera from naturally immunized primates to recognize broadly reactive interspecies antigenic determinants shared by the major structural proteins of type D oncornaviruses, and they suggest possible horizontal transmission of MPMV among rhesus monkeys. Although sera from a number of squirrel monkeys contained antibody to SMRV p35, the possibility that this latter reactivity was due to endogenous virus activation rather than horizontal transmission cannot be ruled out.

Animals↗

Bovine leukemia virus: an exogenous RNA oncogenic virus?

Short term cultures of bovine leukemic lymphocytes release virus particles with biochemical properties of RNA oncogenic viruses. These particles, tentatively called Bovine Leukemia Virus (BLV) have a high molecular weight-reverse transcriptase complex and a density averaging 1.155 g/ml in sucrose solutions. Molecular hybridizations between BLV-3H cDNA and several viral RNAs show that BLV is not related to Mason-Pfizer Monkey Virus (MPMV) Simian Sarcoma Associated Virus (SSV-1) Feline Leukemia Virus (FeLV) or Avian Myeloblastosis Virus (AMV). Rauscher Leukemia Virus (RLV) exhibits a slight but reproducible relatednesse to BLV. The high preference of BLV reverse transcriptase for Mg++ as the divalent cation suggests that BLV might be an atypical mammalian leukemogenic type C virus. Hybridization studies using BLV 3H cDNA as a probe suggest that the DNA of bovine leukemic cells contains viral sequences that cannot be detected in normal bovine DNA.

Animals↗

Bovine leukemia virus: an exogenous RNA oncogenic virus.

Short-term cultures of bovine leukemic lymphocytes release virus particles with biochemical properties of RNA oncogenic viruses. These particles, tentatively called bovine leukemia virus (BLV), have a high molecular weight RNA-reverse transcriptase complex and a density of 1.155 g/ml in sucrose solutions. Molecular hybridizations between BLV/[3H]cDNA and several viral RNAs show that BLV is not related to Mason-Pfizer monkey virus, simian sarcoma associated virus, feline leukemia virus, or avian myeloblastosis virus. These results were confirmed by hybridization between BLV 70S RNA and [3H]cDNA synthesized in the various viruses tested. The high preference of BLV reverse transciptase for Mg++ as the divalent cation suggests that BLV might be an atypical mammalian leukemogenic "type C" virus. DNA-DNA hybridization studies using BLV [3H]cDNA as a probe strongly suggest that the DNA of bovine leukemic cells contains viral sequences that cannot be detected in normal bovine DNA.

Animals↗

The use of trypan blue for counterstaining in the Sepharose bead immunofluorescence test. The application of the test for the demonstration of primate retrovirus-specific antibodies and antigens.

The Sepharose bead immunoflurorescence test was performed by counterstaining the beads with trypan blue. This results in a red staining of negative beads which allows an easy distinction from positive green-fluorescent beads. Sepharose beads conjugated with viral proteins or antiviral antibodies were used to demonstrate Mason-Pfizer monkey virus (MPMW)- and simian sarcoma virus (SSV) - specific antigens or antibodies. The test shows a high sensitivity and specificity and needs a small amount of material.

Antibodies, Viral↗

[Chronic infection caused by the simian Mason-Pfizer virus in primate cell cultures].

Cell cultures of monkey prepuce (Rhfs) and African green monkey kidney (BSC-1) were infected once with simian Mason-Pfizer virus (MPV) and virus expression in the course of establishment of chronic infection was studied. The productive infection was characterized by changes in the cell metabolism (DNA synthesis increased 2-3-fold as early as the "zero" passage), the appearance of gs-antigen, formation of virions of type D and high activity of RNA-dependent DNA-polymerase. Multinuclear giant cells appeared only in the infected Rhfs cell culture most sensitive to MPV. In human embryo kidney culture (HEK) productive infection was also established however, HEK cultures did not survive after 3-4 passages. No signs of transformation could be found in any of the cultures studied.

Animals↗

Molecular diversity among five different endogenous primate retroviruses.

Genetically transmitted retroviruses of Old and New World monkeys include type C viruses isolated from baboons (M7), macaque (MAC-1), and owl monkeys (OMC-1) and type D viruses from langurs (PO-1-Lu) and squirrel monkeys (SMRV, M534). Each of these isolates is unrelated to the others by nucleic acid hybridization criteria and contains a unique array of virion-associated proteins which can be resolved by agarose gel filtration and polyacrylamide gel electrophoresis under denaturing conditions. The major structural protein of each virus has a distinct primary structure, as determined by two-dimensional tryptic peptide analysis, and is antigenically different from the others. The major virion phosphoproteins of endogenous primate type C viruses (pp15) are also different from those of type D viruses (pp13-pp14). Immunological and structural analyses show that the endogenous langur virus and the horizontally transmitted Mason-Pfizer virus of rhesus monkeys are closely related to one another, consistent with the sequence homology detected in their RNA genomes. Although certain radioimmunoassays detect interspecies antigenic determinants common to either the p30 or gp70 proteins of some of these viruses, no one assay has yet been designed which can detect all groups of endogenous primate retroviridae. The data lead to the conclusion that primates contain a minimum of three different sets of genetically transmitted type C and type D retroviral genes.

Animals↗

Use of molecular hybridization to detect type D retrovirus markers in rhesus placentas and other tissues.

We have shown previously that approximately 20% of the Mason-Pfizer virus (MPV) genome is present as endogenous provirus in rhesus monkeys. We report here that several full-term rhesus placentas examined contain additional MPV proviral sequences in their DNA. Competitive molecular hybridization experiments demonstrated that some of these placentas also contain RNA complementary to the entire MPV 60 to 70S RNA genome. Examination of internal organs of rhesus monkeys captured in the wild also revealed the presence of additional MPV proviral sequences and expression of MPV RNA in some tissues. These results provide further evidence that MPV is being transmitted via a non-germ line mechanism in the rhesus population and now demonstrate the placenta as a good source for the identification of retrovirus transcriptional products and proviral DNA.

Animals↗

The use of standard and relaxed hybridization conditions to detect two classes of sequences related to type-D retroviruses in the DNAs of primates.

Using standard hybridization conditions (68 degrees C and 0.4 M sodium phosphate) and assaying for RNAase-resistant RNA . DNA duplexes in the presence of 2 X SSC (1 X SSC is 0.15 M NaCl and 0.015 M sodium citrate), sequences representing approx. 20% of the Mason-Pfizer virus (MPV) genome have previously been shown to be endogenous in DNAs of all Old World monkeys examined (Drohan, W., Colcher, D., Schochetman, G. and Schlom, J. (1977) J. Virol. 23, 36--43). We now report that titration of both the temperature at which hybridizations are carried out and the Na+ concentration at which 125I-labeled MPV RNA . DNA hybrids are scored, reveals a second class of sequences related to the MPV genome in the DNAs of primates. These MPV-related sequences, which are similar to an additional 40% of the MPV genome, are detected when the temperature of hybridization is reduced to 54 degrees C and when the resulting 125I-labeled RNA . DNA duplexes are scored for RNAase resistance in 8 X SSC. These sequences are found in the DNAs of all Old World monkeys examined, and the Tm values of the hybrid duplexes are approx. 6--7 degrees C lower than those of the hybrid duplexes formed using standard conditions. These studies further demonstrate the wide distribution of type-D retrovirus sequences in primates.

Animals↗

Squirrel monkey retrovirus: an endogenous virus of a new world primate.

Squirrel monkey retrovirus (SMRV) was isolated by cocultivation of squirrel monkey lung cells with canine cells. 3H-labeled 60-70S SMRV RNA, isolated from virus grown in canine cells, hybridized to the same extent and to the same Cot1/2 value to the DNA of all tissues of all squirrel monkeys tested; Cot1/2 values show that SMRV proviral sequences are present in the low repetitive range. No SMRV proviral sequences were detected in tissues from a variety of other New World monkeys, Old World monkeys, or apes. Murine, feline, bovine, and canine cells also contain no detectable SMRV proviral sequences. Competitive molecular hybridization studies revealed no detectable sequence homology between the 60-70S RNAs of SMRV and Mason-Pfizer virus (MPV). The virion-associated DNA polymerase of SMRV is similar to that of MPV in that it has a molecular weight of approximately 80,000 and prefers magnesium as a divalent cation using oligo(dG)-poly(rC) as primer-template. The virion-associated DNA polymerase of SMRV can be clearly distinguished from those of MPV and the mouse mammary tumor viruses, however, by its preference for manganese as a divalent cation in the presence of high salt.

Animals↗