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Biomedical subjects

R Risser

Publications and source records attributed to R Risser.

At least 55 records · Page 3Linked to original sources
PubMed1990

Characterization of the fusion domain of the human immunodeficiency virus type 1 envelope glycoprotein gp41.

The human immunodeficiency virus transmembrane glycoprotein gp41 has at its amino terminus a strongly hydrophobic stretch of 28 amino acids flanked by a highly conserved series of polar amino acids. To investigate the role in syncytium formation of the hydrophobic amino terminus of gp41 and the polar border of this hydrophobic region, we introduced eight single-amino acid substitutions and one double-amino acid substitution in the amino-terminal 31 amino acids of gp41. The mutant envelope glycoproteins were expressed from two distinct human immunodeficiency virus type 1 envelope glycoprotein expression vectors; the effects of the mutations on syncytium formation, envelope glycoprotein transport, secretion, and CD4 receptor-binding were analyzed. Results showed that polar substitutions throughout the hydrophobic amino terminus of gp41 greatly reduced or blocked syncytium formation mediated by the human immunodeficiency virus type 1 envelope glycoproteins, as did nonconservative mutations in the polar border of the hydrophobic amino terminus. Mutations at gp41 amino acids 15, 26, and 29 also significantly increased the extent of gp120 secretion into the extracellular medium. None of the mutations detectably affected envelope glycoprotein processing or envelope glycoprotein binding to CD4.

Amino Acid Sequence↗
PubMed1990

Conservation of function of Drosophila melanogaster abl and murine v-abl proteins in transformation of mammalian cells.

The Drosophila melanogaster abl and the murine v-abl genes encode tyrosine protein kinases (TPKs) whose amino acid sequences are highly conserved. To assess functional conservation between the two gene products, we constructed Drosophila abl/v-abl-chimeric Abelson murine leukemia viruses. In these chimeric Abelson murine leukemia viruses, the TPK and carboxy-terminal regions of v-abl were replaced with the corresponding regions of D. melanogaster abl. The chimeric Abelson murine leukemia viruses were able to mediate morphological and oncogenic transformation of NIH 3T3 cells and were able to abrogate the interleukin-3 dependence of a lymphoid cell line. We also found that a virus that contained both TPK and carboxy-terminal Drosophila abl regions had no in vitro transforming activity for primary bone marrow cells and lacked the ability to induce tumors in susceptible mice. A virus that replaced only a portion of the v-abl TPK region with that of Drosophila abl had low activity in in vitro bone marrow transformation and tumorigenesis assays. These results indicate that the transforming functions of abl TPKs are only partially conserved through evolution. These results also imply that the TPK region of v-abl is a major determinant of its efficient lymphoid cell-transforming activity.

Abelson murine leukemia virus↗
PubMed1990

Adenosquamous carcinoma of the cervix: prognosis in stage IB.

The outcome of women with adenosquamous carcinoma of the cervix has been a controversial issue. To assess the prognostic significance of this cell type, the medical records of 29 patients with stage IB primary adenosquamous carcinoma of the cervix were reviewed. Twenty-six had cervical tumors measuring 3 cm or less. With a median follow-up of 60 months, the overall disease-free 5-year survival was 85%. Pelvic lymph node metastases were found in three of 19 patients (16%) with stage IB, and periaortic lymph nodes were negative in eight. A log-likelihood chi 2 test comparing lymphatic spread, recurrence rate, and 5-year disease-free survival in patients with stage IB disease and tumors less than 3 cm versus a similar group of 58 patients with pure squamous cell carcinoma of equivalent stage and comparable size treated during the same time disclosed no significant differences in any of the indicators analyzed. We conclude that stage IB adenosquamous carcinoma of the cervix does not differ in its metastatic potential or outcome from the more common pure squamous cell carcinoma.

Adenocarcinoma↗
PubMed1989

Mutational analysis of the cleavage sequence of the human immunodeficiency virus type 1 envelope glycoprotein precursor gp160.

The envelope glycoproteins of the human immunodeficiency virus (HIV) type 1 are synthesized as a precursor molecule, gp160, which is cleaved to generate the two mature envelope glycoproteins, gp120 and gp41. The cleavage reaction, which is mediated by a host protease, occurs at a sequence highly conserved in retroviral envelope glycoprotein precursors. We have investigated the sequence requirements for this cleavage reaction by introducing four single-amino-acid changes into the glutamic acid-lysine-arginine sequence immediately amino terminal to the site of cleavage. We have also examined the effects of these mutations on the syncytium formation induced by HIV envelope glycoproteins. Our results indicate that a glutamic acid to glycine change at gp120 amino acid 516, a lysine to isoleucine change at amino acid 517, and an arginine to lysine change at amino acid 518 affect neither gp160 cleavage nor syncytium formation. The results obtained with the arginine to lysine change at amino acid 518 differ significantly from the results obtained with the same mutation at the envelope precursor cleavage site of a murine leukemia virus (E. O. Freed, and R. Risser, J. Virol. 61:2852-2856, 1987). An arginine to threonine mutation at gp120 amino acid 518, the terminal residue of gp120, abolishes both gp160 cleavage and syncytium formation. These findings demonstrate that despite its highly conserved nature, the basic pair of amino acids at the site of gp160 cleavage is not absolutely required for proper envelope glycoprotein processing. This report also supports the idea that cleavage of gp160 is required for activation of the HIV envelope fusion function.

Fluorescent Antibody Technique↗
PubMed1989

Multiple steps are required for the induction of tumors by Abelson murine leukemia virus.

Helper virus-free Abelson murine leukemia virus (A-MuLV) was used to induce monoclonal pre-B-cell tumors in mice. The clonality, patterns of immunoglobulin heavy-chain gene rearrangement, tumorigenicity, and v-abl oncogene expression in individual preleukemic and leukemic colonies were compared. Our results indicate that A-MuLV preleukemic cells with low or undetectable tumorigenic potential give rise to leukemic cells with high tumorigenic potential by a process of subclone selection. The levels of v-abl oncogene product in preleukemic and leukemic cell populations were not significantly different. These results suggest that an additional event(s) unrelated to the level of the v-abl protein product is required for A-MuLV-transformed cells to become fully malignant.

Abelson murine leukemia virus↗
PubMed1989

Abrogation of factor-dependence in two IL-3-dependent cell lines can occur by two distinct mechanisms.

We have investigated the mechanisms of abrogation of factor-dependence of lymphoid (FL5.12) and myeloid (FDC-P1) interleukin-3 (IL-3)-dependent cell lines following infection of the cells with retroviruses that encode different oncogenes and immediate selection of IL-3-independent growth. As previously reported by others, Abelson-MuLV (Ab-MuLV) transforms both cell lines to factor-independence, and the transformed cells express the viral oncogene, do not produce IL-3, and are tumorigenic. In contrast, IL-3-independent lymphoid and myeloid cell lines recovered after infection with retroviruses that encode v-src or v-fms oncogenes lacked the v-src or v-fms provirus. Three of seven IL-3-independent transformants were rearranged at the IL-3 locus, produced IL-3-specific mRNA, secreted IL-3 into the culture medium, and grew better in the presence of IL-3. These IL-3-independent transformants produced tumors upon inoculation into nude mice, albeit with longer latent periods than Ab-MuLV transformants. The expression of hematopoietic cell markers was compared in parental and IL-3-independent transformants by immunofluorescence and analysis of mRNA levels. Significant elevation of LFA-1 and MHC class I molecules and loss of expression of IL-2-receptor molecules was detected in most IL-3-independent transformants. These differences in antigen expression indicate that additional phenotypic changes accompany transformation to factor-independence.

Animals↗
PubMed1988

Regulation of expression of T cell gamma chain, L3T4 and Ly-2 messages in Abelson/Moloney virus-transformed T cell lines.

Recent results have suggested that T cells may exist in two distinct pathways, one expressing alpha and beta chain of the T cell receptor genes with either or both of the cell surface markers CD4 and CD8, while the other is negative for these cell surface markers and expresses the T cell-specific gamma chain genes. The relationship between these two pathways is not known. In this study, we have examined a series of either Abelson virus or Moloney virus-derived T cell lines for their expression of these T cell receptor and cell surface marker genes. Results indicate that the Abelson T cell lines do not express the cell surface markers CD4 and CD8, but express relatively high levels of gamma chain transcripts. After culture of these cell lines with the phorbol ester phorbol myristate acetate and interleukin 2, a down-regulation of these gamma chain transcripts can be observed. More interestingly, we found that the Moloney virus-derived T cell lines, which express the cell surface markers CD4 and/CD8, contain high levels of alpha and beta chain T cell receptor transcripts but little or no gamma transcripts even though they have rearranged these latter genes. The gamma transcripts, however, can be induced to high levels after culture with phorbol myristate acetate and interleukin 2. In the process, the cell surface markers CD4 and CD8 and their transcripts were dramatically down-regulated resulting in cells with high levels of gamma chain transcripts and a CD4-CD8- phenotype. The regulation of expression of these genes is reversible. Taken together, these results indicate that the T cell receptor gamma chain genes and those of the cell surface markers CD4 CD8 can be regulated in vitro by external factors and it opens up the possibility of studying the regulatory sequences associated with these genes.

Abelson murine leukemia virus↗
PubMed1988

Mechanism of interaction between endogenous ecotropic murine leukemia viruses in (BALB/c X C57BL/6) hybrid cells.

The germline ecotropic murine leukemia (MuLV) proviruses of BALB/c and C57BL/6 (B6) mice were analyzed to determine the molecular basis of low virus expression in these mouse strains and to determine the mechanism of interaction of these two proviruses. Previous work had demonstrated that the BALB/c endogenous ecotropic provirus was infectious but unable to induce XC cell syncytia formation, and that induced (BALB/c X B6) hybrid cells expressed 10- to 50-fold more XC syncytia than induced parental cells. Two independently isolated DNA clones of the B6 endogenous ecotropic provirus were noninfectious following transfection into cells, and cell lines that expressed this viral genome produced noninfectious MuLV. Nucleotide sequencing of the mutant region of the B6 provirus indicated that the defective nature of this provirus resulted from an amino acid substitution of proline for alanine in the central portion of reverse transcriptase. From the analysis of the virus produced by induced hybrid cells, and the patterns of steady-state viral RNA in induced cells, we propose that the enhanced XC cell syncytia formation observed in hybrid cells is due to trans-complementation of viral proteins and not viral recombination or trans-activation of viral genome expression.

Animals↗
PubMed1988

Substitution of the LTR of Abelson murine leukemia virus does not alter the cell type of virally induced tumors.

The long terminal repeat (LTR) of the pre-B cell tropic Abelson murine leukemia virus (A-MuLV) was replaced with the LTR of the erythrotropic Friend MuLV or with the LTR of the erythropic/fibrotropic Harvey murine sarcoma virus (Ha-MuSV) to generate the viruses F-ABL and H-ABL, respectively. The parental A-MuLV and the recombinant viruses induced pre-B cell lymphomas in susceptible mice with similar frequencies. Recombinant virus-induced tumor DNAs were analysed by nucleic acid hybridization and were shown to contain the appropriate recombinant provirus. F-ABL was 100-1000 fold less efficient than A-MuLV or H-ABL in the in vitro transformation of primary bone marrow cells, as detected by lymphoid colony formation in agarose. To compare the level of transcription initiated from the different viral LTRs, we investigated the ability of the U3 region of these retroviral LTRs to promote transcription in a battery of cell lines using the chloramphenicol acetyl-transferase (CAT) assay, and with some exceptions we found the following hierarchy of activities: Ha-MusSV greater than or equal to M-MuLV greater than A-MuLV greater than F-MuLV, regardless of the cell line transfected. These results indicate that the LTR is not a determinant of the pre-B cell disease specificity of A-MuLV, and suggest that this specificity resides in the v-abl oncogene. Also, our results suggest that a threshold amount of the v-abl protein product is necessary for in vitro transformation, and this level of expression may be different from the level selected during in vivo tumorigenesis.

Abelson murine leukemia virus↗
PubMed1987

Nucleotide conservation of endogenous ecotropic murine leukemia proviruses in inbred mice: implications for viral origin and dispersal.

Nucleotide sequence analysis of the ecotropic murine leukemia proviruses of AKR, BALB/c, and C57BL/6 mice indicated that these viral genomes differ from each other in less than 0.5% of their sequenced nucleotides, whereas they differ from the laboratory Moloney, Friend, or RadLV viruses or a partial ecotropic provirus found in wild mice by 8-22% of their sequenced nucleotides. The limited variation of endogenous ecotropic proviruses found in these common mouse strains indicates that few cycles of virus replication separated the introduction of the ecotropic endogenous retroviruses into the germlines of the progenitors of these now divergent mouse strains, and is consistent with the hypothesis that these common inbred strains were derived from a pool of very few mice, at least one of which was infected with an ecotropic murine leukemia virus. Ecotropic germline proviruses now found in common inbred mice most likely derive from germline reintegrations of the viral progeny of that initial single infection.

Animals↗
PubMed1987

Cell transformation and tumor induction by Abelson murine leukemia virus in the absence of helper virus.

We investigated the role of the Moloney helper virus, Moloney murine leukemia virus (Mo-MuLV), in cell transformation and tumor induction by the defective Abelson murine leukemia virus (Ab-MuLV). A molecular clone of Ab-MuLV (P160 strain) was transfected into the psi 2 packaging cell line, and helper virus-free Ab-MuLV (psi 2) was harvested from the supernatant medium. Ab-MuLV (psi 2) was as efficient as helper virus-containing Ab-MuLV (Mo-MuLV) in the transformation of primary bone marrow cells in vitro. Inoculation of weanling BALB/c mice with Ab-MuLV (psi 2) induced nonthymic pre-B-cell lymphomas with high efficiency and short latency (28 days). Adult BALB/c mice were less sensitive to tumor induction by a factor of 100. Ab-MuLV (psi 2) did not induce tumors in weanling C57BL/6 mice, unlike Ab-MuLV (Mo-MuLV). Examination of the proviral integration pattern in Ab-MuLV (psi 2)-induced tumor cell DNA revealed that each of the tumors contained a single integrated provirus. Immunoprecipitation of viral-encoded proteins in helper virus-free tumor cell lines detected the P160 Ab-MuLV-transforming protein; however, no trace of the gag, pol, and env helper virus-encoded proteins was found. Our results indicate that integration and expression of a single Ab-MuLV genome is sufficient for efficient transformation of primary bone marrow cells by Ab-MuLV in vitro and tumor induction in susceptible mice. However, the helper virus may contribute to tumor induction in weanling resistant mice.

Animals↗
PubMed1987

Germ line integration of a murine leukemia provirus into a retroviruslike sequence.

Nucleotide sequence analysis of the cellular sequences flanking the integrated ecotropic (mouse-infectious) murine leukemia provirus of BALB/c mice indicated that the murine leukemia provirus is integrated in opposing transcriptional orientation within a solo long terminal repeat (LTR) of the VL30 family of endogenous retrovirus-related sequences. To quantify the effect of this integration event on the ability of the ecotropic provirus to be expressed, we constructed recombinant molecules that carried the chloramphenicol acetyltransferase (cat) gene and various viral LTRs and determined the CAT activity induced by these constructs after transfection of NIH 3T3 cells. Our results indicate that the BALB/c ecotropic LTR is about 10-fold more active than the VL30 LTR. The presence of the VL30 LTR did not affect the transcriptional activity of the ecotropic LTR in the context of the integration event. Our results also indicate that the LTRs of the BALB/c provirus are less transcriptionally active than are the proviral LTRs of AKR murine leukemia virus and the Harvey murine sarcoma virus.

Animals↗
PubMed1987

Clonal dominance and progression in Abelson murine leukemia virus lymphomagenesis.

We examined the clonality of tumors induced by an acutely transforming retrovirus which carries a single oncogene. Contrary to our expectation, tumors induced by the Abelson murine leukemia virus (A-MuLV) showed one to four major proviral integration events. To further investigate the process by which clonality was established, we analyzed the number of cells infected and transformed by A-MuLV at various times after in vivo infection. At the midpoint of tumor latency (14 days postinfection), we found that infection of total bone marrow cells by A-MuLV was efficient and polyclonal. However, only a minority of these infected cells were transformed as assayed in cell culture, and clonal dominance had already been established in this transformed cell population. Examination of the in vitro growth properties of transformed cells recovered from preleukemic and leukemic mice indicated that preleukemic cells had lower cloning efficiencies than primary tumor cells. Our results suggest that the rate-limiting step in this system of lymphomagenesis is the initial transformation of bone marrow target cells and that these cells undergo subsequent changes in cloning ability during the course of the disease that lead to an autonomous neoplastic state.

Abelson murine leukemia virus↗