Bijay K. Pal

Structural analysis of the major envelope glycoprotein (gp70) of the amphotropic and ecotropic type C viruses of wild mice

Abstract Virus-associated and free serum gp70s from wild mice ( Mus musculus ) were isolated by immunoaffinity column chromatography and were analyzed by two-dimensional tryptic peptide mapping procedures. Our results show that the gp70s of wild mouse ecotropic and amphotropic MuLVs are structurally divergent. The gp70s of wild mouse ecotropic viruses are more similar in structure to the Gross-AKR MuLV subgroup than to the FMR subgroup. The gp70s of the amphotropic wild mouse viruses are highly conserved in structure and are closely related to those of one group of xenotropic MuLVs. Non-virion-associated free serum gp70 of wild mice is homologous to that of inbred laboratory mouse strains.

Subviral components of a wild mouse embryo-derived type C oncornavirus

Abstract A wild mouse embryo-derived nontransforming type C virus, WM-1504E, which showed in vivo neurotropic and lymphomagenic activities, was analyzed for biochemical and antigenic characterizations. The virions contained two major species of RNA, 70 S and 4 S. The 70 S RNA was heat dissociated into a heterogeneous species with peak distributions at 35 S, 18 S, and 4 S regions. The gel filtration pattern in guanidine-hydrochloride and neutralization by antibody against MuLV polymerase of the virion associated reverse transcriptase resembled those of MuLV enzyme. Virions contained six major protein groups as detected by agarose gel filtration in guanidine-hydrochloride. The two largest proteins were glycoproteins, and the next largest protein contained the group-specific antigenic determinants of murine type C oncornaviruses. The approximate molecular weights of the major proteins as determined by SDS-polyacrylamide gel electrophoresis were: 100,000 (and 11,000), 76,000, 29,000, 16,000, 13,500, and 11,000 daltons. The last group also contained a polypeptide of 10,000 daltons. These values correspond closely to those of MuLV (Moloney) proteins. It is concluded that WM-1504E virus represents a murine type C oncornavirus.

Genetic relationship of wild mouse amphotropic virus to murine ecotropic and xenotropic viruses

Abstract The genomic relationship between amphotropic and ecotropic wild mouse (Mus musculus) leukemia viruses (MuLV) and between these viruses and the ecotropic and xenotropic MuLVs of inbred mice was determined by DNA-RNA hybridization using viral complementary DNA and polyA-containing 70S viral RNA. The results indicate that (a) the genomes of individual cloned amphotropic and ecotropic wild mouse virus isolates are closely related and (b) these nucleotide sequences are also related to, yet distinguishable from, the genomes of prototype ecotropic (Rauscher, Gross-AKR) and xenotropic (AT-124, NZB, AKR, Balb/c) MuLVs of inbred mice.

RNA tumor virus phosphoproteins: subvirion location of the multiple phosphorylated species.

Abstract An analysis of core particles of 1.23 g/cm 3 density of the endogenous feline type C virus (RD-114) showed the presence of all of the major low molecular weight structural proteins, namely p30, p12, p10, and the phosphoprotein pp15, in relative proportions similar to those detected in intact RD-114 virions. The protein kinase activity of the virus was also found to be associated with the core. A comparison of the content of multiple phosphorylated species of the pp15 derived from the core with that isolated from the whole virion demonstrated that all major phosphorylated species of RD-114 pp15 were located in the core structure.

Endogenous type C RNA virus of mink (Mustela vison).

Abstract A type C RNA virus was isolated from mink lung cell line (American Type Culture Collection No. CCL 64) which had been cocultivated with 5-bromodeoxyuridine (BUDR)-treated mouse spleen cells. The virus has type C RNA virus morphology as demonstrated by electron microscopy. The complement fixation and immunofluorescent tests performed with mouse anti-p30 antisera show a distinctive difference between mink and mouse type C viruses. Complement fixation tests also indicate that mink type C virus is antigenically different from rat, feline leukemia, feline endogenous (RD-114), baboon, and woolly monkey type C viruses. The virus propagates in cells of mouse, rat, cat, sheep, dog, and human origin, but not in bovine (MDBK) or simian (BSC-1) cells. The infection of rabbit (SIRC) cells and cells of virus origin (mink lung) was followed by delayed and low-titer polymerase release in tissue culture media. The virus sediments in sucrose density gradients as a broad band of densities, 1.13–1.17 g/ml, and contains 70 and 4S RNA. The protein profile is similar to that observed in other mammalian type C viruses. The DNA complementary to the poly(A)-containing virion RNA hybridized to a high degree (72%) with the RNA from virus-producing mink lung cells but not with the RNA from mouse cell lines or uninfected mink lung cell line. The nucleotide sequences homologous to mink viral cDNA were found in mink cell DNA from both virus-producing and nonproducing cells, but not in the DNA of mouse, rat, or feline origin. The virus here described therefore represents an endogenous mink type C virus.

RNA tumor virus phosphoproteins: phosphorylation of precursor and processed polypeptides.

Abstract Monospecific antisera made against the 30,000 molecular weight major internal polypeptide (p30) and the 12,000 molecular weight phosphorylated polypeptide (pp12) of a wild mouse type C oncovirus were used to immunoprecipitate precursor polypeptides from extracts of isotopically labeled cells infected with the oncovirus. Analysis of the immunoprecipitates by SDS-polyacrylamide gel electrophoresis led to the detection of several precursor polypeptide (Pr) species containing the determinants of both p30 and pp12. These species, namely Pr>100, Pr100, Pr77, Pr62, and Pr50, were all found to be phosphorylated in pulse experiments. The polypeptide pp12 was, however, the major phosphorylated species immunoprecipitated by anti-pp12 sera in pulse and chase experiments. These data and a relatively high degree of phosphorylation in the processed pp12 suggested that phosphorylation of oncovirus protein is initiated at the polyprotein level and the phosphoprotein moiety is further phosphorylated subsequent to processing.

RNA tumour virus phosphoproteins: evidence for virus specificity of phosphorylation.

Summary The purified 12000 dalton (p12) phosphoprotein of Rauscher (R) and wild mouse (WM) strains of murine leukaemia virus (MuLV) was analysed for the distribution patterns of its variously charged molecular species by urea-poly-acrylamide gradient gel electrophoresis. The distribution patterns of the p12 of two different field isolates of WM viruses, 292 and 1504, and the mouse-tropic and amphotropic clonal sub-populations of 15-4 field isolate were very similar but different from that of MuLV-R. A unique characteristic of the p12 of the WM isolates is the presence of two major apparently non-phosphorylated species in approximately constant proportions relative to the phosphorylated species. Similar studies on the p12 of the same virus (MuLV-R or WM viruses) grown in different host cells showed that the patterns of phosphorylated and non-phosphorylated species are virus-specific and independent of the cell lines of propagation. These analyses and their comparison with urea-gel patterns of the phosphoproteins of other mammalian type C viruses indicated that the number and relative proportion of the variously phosphorylated and non-phosphorylated species are predetermined for a virus. Therefore, the virus must have the genetic information for the phosphoprotein as well as other necessary genetic information which functions, perhaps in conjunction with appropriate cellular factors, in regulating the specific proportions of these multiple species. Possible biological significance of the variously charged molecular species in the phosphoprotein of RNA tumour viruses is discussed.

Major phosphoprotein of avian sarcoma virus: Peptide analysis of the variously charged species

Abstract The 19,000 dalton phosphoprotein (pp19) of avian sarcoma virus strain B77 isolated by guanidine-agarose chromatography was further resolved into multiple charged species by DEAE-cellulose chromatography. A comparison of the tryptic peptide maps of 125I-iodinated lowly phosphorylated and highly phosphorylated species of the pp19 suggested that the same polypeptide occurs in the virion in different phosphorylated states.

Structural polypeptides of primate derived type C RNA tumor viruses.

Abstract Proteins of gibbon ape lymphosarcoma virus (GaLV) and woolly monkey sarcoma virus, type 1, together with its associated virus ( SSV-1 SSAV-1 ) were analyzed by guanidine-agarose chromatography and the separation patterns were compared with those of mouse and feline type C viruses. GaLV contained five major proteins, including two glycoproteins, whereas lower mammalian viruses contained six major proteins, including two glycoproteins. The molecular weights of the five GaLV proteins closely resembled the molecular weights of the five equivalent lower mammalian viral proteins. SSV-1 SSAV-1 showed a separation pattern similar to GaLV except it contained a low but detectable amount of an additional glycoprotein. Both GaLV and SSV-1 SSAV-1 were deficient in a protein of molecular weight about 15,000 daltons which is found in all known type C viruses of avian, reptilian and lower mammalian species.

AKvr-1, A DOMINANT MURINE LEUKEMIA VIRUS RESTRICTION GENE, SEGREGATES IN LEUKEMIA-PRONE WILD MICE

ABSTRACT Leukemia-prone wild mice ( Mus musculus domesticus ) from a squab farm near Lake Casitas (LC) in southern California are polymorphic for a restriction gene, named Akvr-1, that suppresses AKR ecotropic virus. The restriction allele (Akvr-1R) is dominant and exhibits 100% penetrance in prevention of viremia of AKR endogenous retrovirus and of virus-associated lymphoma in (LCRR X AKR) F1 hybrids. Ecotropic and xenotropic AKR virus production is also suppressed in thymus, spleen and bone marrow of older resistant F1 hybrids. The restriction phenotype segregates as a single Mendelian locus in F2 and backcrosses to AKR mice. Akvr-1R likewise is effective in restriction of NB-tropic Moloney and Friend MuLVs in vivo but fails to restrict expression or pathogenesis of LC-derived amphotropic retrovirus. This gene apparently exerts a similar MuLV-suppressive effect in vitro upon various cell types. Akvr-1 does not map near to certain retroviral gene loci including Fv-1, Fv-2 and Akv-1. It may be related to the Fv-4 locus described in Japanese mice.