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E Scolnick

Publications and source records attributed to E Scolnick.

At least 19 recordsLinked to original sources
PubMed1983

Envelope gene of the Friend spleen focus-forming virus: deletion and insertions in 3' gp70/p15E-encoding region have resulted in unique features in the primary structure of its protein product.

A nucleotide sequence was determined for the envelope (env) gene of the polycythemia-inducing strain of the acute leukemia-inducing Friend spleen focus-forming virus (SFFV) and from this the amino acid sequence of its gene product, gp52, was deduced. All major elements of the gene were found to be related to genes of other retroviruses that code for functional glycoproteins. Although the carboxyl terminus of gp52 is encoded by sequences highly related to sequences in its putative parent, ecotropic Friend murine leukemia virus, the majority of the protein (69%), including the amino terminus, is encoded by dualtropic virus-like sequences. Nucleotide sequence comparisons suggest that the nonecotropic region may be more closely related to the 5' substitution in dualtropic mink cell focus-inducing viruses that it is to the 5' end of xenotropic virus env genes. A large deletion and two unique insertions have been located in the env gene of polycythemia-inducing SFFV and may account for some of the unusual structural characteristics, aberrant processing, and pathogenic properties of gp52. As a consequence of the deletion, amino-terminal gp70 and carboxyl-terminal p15E-encoding sequences are juxtaposed and it appears that translation from the p15E region, 3' to the deletion, continues in the standard reading frame used by other retroviruses. Insertions of six base pairs and one base pair at the very 3' end of the gp52-encoding region results in a SFFV-unique amino acid sequence and a premature termination codon.

Amino Acid Sequence↗
PubMed1983

Altered growth and differentiation of cultured mouse epidermal cells infected with oncogenic retrovirus: contrasting effects of viruses and chemicals.

Mouse epidermal basal cells can be cultured in medium with a Ca2+ concentration below 0.1 mM and induced to terminally differentiate in medium with higher Ca2+. Infection of basal cells with Kirsten or Harvey sarcoma virus causes a marked stimulation of basal cell proliferation. Challenge of infected basal cell cultures with 0.5 mM Ca2+ medium indicates that terminal differentiation is altered, since infected cultures persist in this medium for long periods without stratifying and sloughing from the culture dish. Infected cultures express much higher levels of protein with a molecular weight of 21,000 than controls, and studies with temperature-sensitive mutants indicate that persistent function of protein with a molecular weight of 21,000 is required for the observed effects of the sarcoma viruses on terminal differentiation. In response to 0.5 mM Ca2+, the proliferation rate is markedly reduced in both control and virally infected cells. However, the proliferation rate of the sarcoma virus-infected cultures is reduced less in response to elevated levels of calcium. Virus-infected cells as well as controls also demonstrate a rise in the differentiation-associated enzymatic activity of epidermal transglutaminase and will terminally differentiate when the multiplicity of infection is below one. These results suggest that sarcoma virus-infected cells are not blocked completely in their program of terminal differentiation in response to 0.5 mM Ca2+ and differ in this respect from epidermal cells altered by exposure to chemical carcinogens and from malignant keratinocytes. Infection with Kirsten or Harvey sarcoma virus and expression of protein with a molecular weight of 21,000 therefore are closely linked to epidermal proliferation and retard, but do not completely block, the program of terminal differentiation. The effects of ras oncogenes on NIH 3T3 cells are discussed relative to the effects of ras oncogenes observed on the primary mouse epidermal cells in the context of multistep models for naturally occurring epithelial cancers.

Animals↗
PubMed1982

Nucleotide sequence of the p21 transforming protein of Harvey murine sarcoma virus.

Harvey murine sarcoma virus is a retrovirus which transforms cells by means of a single virally encoded protein called p21 has. We have determined the nucleotide sequence of 1.0 kilobase in the 5' half of the viral genome which encompasses the has coding sequences and its associated regulatory signals. The nucleotide sequence has identified the amino acid sequence of two additional overlapping polypeptides which share their reading frames and the carboxyl termini with p21 but which contain additional NH2-terminal amino acids.

Amino Acid Sequence↗
PubMed1981

Friend disease in vitro.

In long-term marrow cultures, hemopoiesis can be maintained for several months, although erythropoiesis is normally suppressed at the most primitive level of development (the erythroid colony-forming cells). Infection of these cultures with a viral complex combining helper-independent murine leukemia virus (F-MuLV) and a spleen focus-forming virus (SFFVp) results in a productive infection of both the replication defective SFFVp and the F-MuLV. After infection, the cultures show a dramatic elevation in the numbers of late erythroid progenitor cells (CFU-E), many of which will grow in the absence of added erythropoietin, and a transient erythropoietin, independent erythropoiesis, including the production of mature, enucleated erythrocytes. Hemopoiesis eventually declines, with no evidence for the generation of Friend tumor cells. When erythropoiesis is induced in the long-term cultures by addition of anemic mouse serum before infection by polycythemia-inducing Friend virus, the generation of erythropoietin-independent CFU-E and erythrocyte formation is followed by the sustained production (greater than 40 wk) of primitive erythroid cells with low spontaneous levels (less than 5%) of hemoglobinization. Although these cells will produce spleen colonies in irradiated mice and can be cloned in soft-gel media, they do not produce autonomous, permanently growing cell lines in vitro, i.e., they retain a dependency upon the marrow-adherent layer for their continued growth. However, following a further passage on a "virgin" marrow environment, permanent cell lines can be established that are able to grow independently of environmental influences. Thus, this system is the first description of a complete in vitro system for the reproducible production and isolation of Friend virus-induced erythroid cell lines.

Anemia↗
PubMed1981

Isolation of transplantable erythroleukemia cells from mice infected with helper-independent Friend murine leukemia virus.

The Friend virus complex contains a helper-independent retrovirus, F-MuLV, and a replication-defective retrovirus, SFFV. Murine erythroleukemia cell lines (MEL) have been isolated previously from the leukemic tissues of mice infected with such stocks of Friend virus complex. Since our laboratory has shown that either F-MuLV or SFFV can induce a lethal erythroproliferative disease, it has been unclear which of the properties of conventional MEL cells were causally related to the infection of these cells by F-MuLV and which properties were related to the infection of these cells by SFFV. In this article, we have focused on the helper-independent component of the Friend virus complex, F-MuLV clone 57, which has recently been purified by molecular cloning. We report a method for the reproducible isolation of a new type of murine erythroleukemia cell that can be isolate from mice infected with F-MuLV clone 57. We have designated these cells as 57 TP-1, indicating their origin from mice infected with F-MuLV clone 57 and their characteristics as transplantable erythroleukemia cells. Upon subcutaneous, intraperitoneal, or intravenous inoculation of these cells, 57 TP-1 cells grow in the hematopoietic organs of the recipient mice. No subcutaneous or intraperitoneal nodules occur. Both F-MuLV and a Friend MCF virus can be detected in these cells, but no evidence to SFFV can be found.

Animals↗
PubMed1981

Molecular and cell biologic aspects of erythropoiesis in long-term bone marrow cultures.

In long-term marrow cultures, proliferation and differentiation of hemopoietic stem cells occurs for several months. Normally, only the most primitive erythroid progenitor cells are produced (the BFU-E). Following treatment with anemic mouse serum (AMS) or normal mouse serum plus erythropoietin, the BFU-E mature into CFU-E, which then go to produce mature nonnucleated red cells. This development is associated with the production of adult type hemoglobin. Furthermore, erythropoiesis and granulopoiesis occur in association with discrete cellular elements of the adherent cell layer in the long-term culture. Following treatment with AMS, erythropoiesis is enhanced while granulopoiesis is depressed, with no apparent competition at the stem cell or progenitor cell level.

Adipose Tissue↗
PubMed1980

Growth of factor-dependent hemopoietic precursor cell lines.

Cell lines have been produced from long-term cultures of mouse bone marrow that require a factor, present in WEHI-3 conditioned medium (CM) or in spleen CM, for their sustained growth. The cell lines were obtained from nonvirus-treated cultures, are nonleukemic, maintain a normal karyotype, and form colonies showing granulocyte maturation when plated in soft agar. Granulocyte/macrophage (GM) colony-stimulating factor is not the inductive moiety involved in the maintenance of proliferation of these cells. It is suggested that the cell lines represent a self-renewing population of cells ancestral to GM colony-forming cells, which may be responding to a hitherto unrecognized regulator.

Animals↗
PubMed1980

Three laboratory strains of spleen focus-forming virus: comparison of their genomes and translational products.

The molecular properties of three laboratory strains of the spleen focus-forming virus were compared. All strains contain genetic sequences related to the env gene of mink cell focus-inducing murine type C leukemia viruses, and each strain codes for a glycoprotein of 50,000 to 52,000 daltons which shares specific immunological properties with the gp70's of mink cell focus-inducing viruses. In contrast to this constancy, gag gene products coded for by these strains vary significantly. The gag and env gene products are synthesized from separate mRNA's, and the mRNA for the env gene product is approximately 18S. Unlike other acute leukemia viruses, which can transform various undifferentiated cells, have large unique sequence cellular gene inserts fused to helper virus gag genes, and have one known genome-length intracellular mRNA, the spleen focus-forming virus transforms only specific hematopoietic stem cells, is an env gene rather than a gag gene recombinant virus, and has a second distinct and smaller class of intracellular mRNA. Our data therefore indicate that the Friend strain of the spleen focus-forming virus is a unique replication-defective acute leukemia virus.

Animals↗
PubMed1980

Subgenomic fragment of molecular cloned Friend murine leukemia virus DNA contains the gene(s) responsible for Friend murine leukemia virus-induced disease.

Friend murine leukemia virus (G-MuLV) is a helper-independent, type C retrovirus isolated from stocks of Friend virus complex (spleen focus-forming virus plus MuLV). In cell culture, F-MuLV has an ecotropic and NB-tropic host range and causes XC cells to fuse. When injected into newborn NIH Swiss mice, F-MuLV produces hepatosplenomegaly, severe anemia, and numerous circulating hematopoietic precursors in the peripheral blood with normal thymus and lymph nodes after 3 to 6 weeks. Recently, we molecularly cloned an 8.5-kilobase pair (kbp) form of F-MuLV DNA from which we could recover the pathogenic F-MuLV virus by DNA transfection of NIH 3T3 cells. From this molecularly cloned F-MuLV DNA, we have now subcloned in pBR322 a 4.1-kbp HindIII fragment which contains in continuity 3.0 kbp from the 3' terminus (env and c region), 0.6 kbp of the terminal repeat sequences, and 0.5 kbp from the 5'terminus of the viral RNA (genome). NIH 3T3 fibroblasts were transfected with this DNA fragment an then infected with the wild mouse amphotropic retrovirus (cl 1504-A). In cell culture, 1504-A is a helper-independent type C virus which has an N-tropic host range and does not cause fusion of XC cells. When injected into newborn NIH Swiss mice, 1504-A does not produce splenomegaly or thymic enlargement in mice held for up to 8 months. The transfection with the F-MuLV fragment and the infection with 1504-A consistently yielded virus preparations that were XC positive. From such virus stocks we were able to isolate both helper-independent and replication-defective XC-positive viruses. The helper-independent virus was shown to be a recombinant virus since it contains a gp70 molecule derived at least in part from F-MuLV and a specific gag precursor derived from 1504-A as determined by radioactive immune precipitation assays. When injected into newborn Swiss mice, the recombinant helper-independent virus caused hepatosplenomegaly in approximately 50% of the mice in 6 to 8 weeks. The histology of the diseased splenic tissue was indistinguishable from that seen in the disease caused by the whole F-MuLV. The replication-defective virus could be pseudotyped with new 1504-A virus, and this viral complex also caused the F-MuLV disease picture when the complex was injected into newborn Swiss mice. We conclude that the genetic information responsible for the pathogenicity of F-MuLV is contained within the 4.1-kbp DNA fragment, which includes env gene sequences, the terminal repeat sequences, and the c region sequences of the F-MuLV genome.

Animals↗
PubMed1978

Type-specific radioimmunoassays for the gp70s of mink cell focus-inducing murine leukemia viruses: expression of a cross-reacting antigen in cells infected with the Friend strain of the spleen focus-forming virus.

We have isolated the gp70 of a helper-independent strain of a Friend mink cell focus-inducing (MCF) virus, Fr-MCF-1. This recombinant virus, like the previously described AKR-MCF viruses, has been shown by both biological and biochemical means to be an envelope gene recombinant between Friend murine leukemia virus (F-MuLV) and a mouse xenotropic virus. Utilizing (125)I- labeled Fr-MCF-1 gp70 and antiserum prepared against an MCF strain of Moloney type-C virus (Mol-MCF(83)), we have developed a radioimmunoassay which detects immunological determinant (s)contained in the gp70s of MCF viruses derived from F-MuLV, Mol-MuLV, and AKR-MuLV. This MCF determinant(s) is not detected in the ecotropic parents of each of these MCF viruses, nor in helper-independent murine xenotropic viruses derived from Swiss or BALB/c mice. A protein partially cross-reactive with the MCF gp70 determinant(s) is detected in a replicating xenotropic virus derived from NZB mice. Utilizing this MCF gp70 specific immunoassay, we can detect a cross-reacting gene product coded for by the Friend strain of the spleen focus-forming virus (SFFV) in rat fibroblasts nonproductively infected with SFFV. The results support earlier molecular hybridization studies which indicated that the genome of SFFV contains genetic information derived from both F-MuLV and xenotropic virus, and that the xenotropic-related sequences in SFFV are highly related to those found in MCF murine type-C viruses.

Antigens, Viral↗
PubMed1976

Long-term persistent vesicular stomatitis virus and rabies virus infection of cells in vitro.

BHK 21 carrier cells persistently infected with VSV Indiana for over 2 years have been shedding generally very low levels of mature infectious virus or mature T particles (averaging less than one-hundredth p.f.u./cell/day) yet most cells are producing virus antigens and are resistant to homologous superinfection. However, large amounts of biologically active T particle RNP can be recovered from cytoplasmic extracts of these carrier cells even at times when they are shedding no detectable infectious virus. This recovered cytoplasmic RNP replicates (with helper B virions) to produce mature T particles, interferes strongly after DEAE dextran-facilitated uptake and, together with B virions, allows the establishment of a persistent carrier state in exposed cells. No 'provirus' DNA copies of the VSV RNA genome are detectable (less than 1/40 copy/cell or I copy per 40 cells) in carrier cells after more than 2 years of persistent infection, and all transfection attempts have failed using DNA from these VSV carriers or DNA from carrier cells persistently infected with some other negative strand RNA viruses (measles, mumps, LCM, influenza, rabies). Infectious viruses shed after more than I year from carrier cells originally infected with wild-type B virions are small plaque mutants showing a slight temperature sensitivity. Cured cell populations can be obtained from the long term VSV carrier culture by cloning in the presence or absence of antiviral antibody.

Animals↗
PubMed1972

In vitro synthesis of DNA complementary to purified rabbit globin mRNA (RNA-dependent DNA polymerase-reticulocyte-hemoglobin-density gradient centrifugation-oligo(dT) primer).

Several properties of the viral RNA-dependent DNA polymerases and of rabbit globin mRNA make it possible to consider synthesis of the globin gene in vitro. These enzymes copy an RNA template using a short sequence of complementary nucleotides as a primer. Furthermore, globin mRNA has a 3'-terminal sequence of adenylic acid residues that make it particularly suitable as a template, since oligo(dT) can be annealed to a specific site on the mRNA. This small primer could phase the DNA polymerase, possibly ensuring that replication is initiated from that end of the globin message. We have used this approach and find that purified mRNA is an efficient template for the polymerase enzyme. The reaction requires the RNA template and the four deoxyribonucleoside triphosphates, and it is markedly stimulated by the addition of oligo(dT). Consistent with the expectation that the oligo(dT) uniquely phases the polymerase at an adenine-rich region in the globin message, oligo(dG), oligo(dC), and oligo(dA) fail to serve as primers. The product has a density intermediate between that of DNA and RNA, and shifts to a lighter DNA density after treatment with base. Further, it is specifically complementary to globin mRNA and sediments slightly faster in an alkaline sucrose gradient than a DNA standard that has a molecular weight of 129,000. The data suggest that a major portion of the DNA product is a sequence of at least 500 bases, about 50 more than would be necessary to encode rabbit globin. The potential usefulness of this interesting product is discussed.

Animals↗
PubMed1970

Peptide chain termination. VI. Purification and site of action of S.

PEPTIDE CHAIN TERMINATION IS A RESULT OF AT LEAST TWO EVENTS: terminator codon recognition and hydrolysis of peptidyl tRNA. A protein factor S, isolated from the supernatant of Escherichia coli B, stimulates fMet release. Factor S lowers the K(m) for terminator trinucleotides without altering the V(max) of release and therefore acts at terminator codon recognition. The S protein differs from initiation factors, elongation factor G, several forms of elongation factor T and release factors. The importance of the 2% Tu content in purified S is unresolved.

Bacterial Proteins↗
PubMed1970

RNA-dependent DNA polymerase activity in five RNA viruses: divalent cation requirements.

The RNA-dependent DNA polymerase(s) in murine leukemia, murine mammary tumor, and avian myeloblastosis viruses require maganese for optimal activity. The transcription of added synthetic polyribonucleotides is greatly enhanced when manganese is used in place of magnesium. A soluble RNA-dependent DNA polymerase activity has been released from murine leukemia particles in the presence of manganese and high detergent concentrations. Two RNA viruses, visna virus of sheep and a primate syncytial virus, not known to have tumor-producing ability, also contain the polymerase.

Adenine Nucleotides↗