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T P Bender

Publications and source records attributed to T P Bender.

29 records · Page 2Linked to original sources
PubMed1987

Differential expression of c-myb mRNA in murine B lymphomas by a block to transcription elongation.

Expression of c-myb proto-oncogene messenger RNA (mRNA) and protein has been detected principally in tumors and in normal tissue of hematopoietic origin. In each hematopoietic lineage examined, expression of the c-myb gene is markedly downregulated during hematopoietic maturation. However, the mechanism by which differential expression of the c-myb gene is regulated is not known. In murine B-lymphoid tumor cell lines, the amount of steady-state c-myb mRNA is 10 to more than 100 times greater in pre-B cell lymphomas than in B cell lymphomas and plasmacytomas. The downregulation of c-myb mRNA correlates with events at the pre-B cell-B cell junction. Differential expression of c-myb mRNA levels detected between a pre-B cell lymphoma and a mature B cell lymphoma is now shown to be mediated by a block to transcription elongation in the first intron of the c-myb locus. In addition, this developmentally regulated difference in transcriptional activity is correlated with alterations in higher order chromatin structure as reflected by changes in the patterns of hypersensitivity to deoxyribonuclease I at the 5' end of the c-myb transcription unit. Regulation of transcription elongation may provide a more sensitive mechanism for rapidly increasing and decreasing mRNA levels in response to external stimuli than regulation of the initiation of transcription.

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PubMed1986

Murine myb protooncogene mRNA: cDNA sequence and evidence for 5' heterogeneity.

We have sequenced two overlapping cDNA clones from a murine pro-B cell library to generate a composite sequence that includes 3413 bases of the murine c-myb mRNA. There is a single long open reading frame, beginning at the first base of this sequence, and continuing from the first methionine codon at nucleotide 265 to a TGA termination codon at nucleotide 2173. The predicted murine translation product contains 636 amino acid residues and is about 71 kDa long, which is in good agreement with the 75-kDa molecular size determined for the avian c-myb protein. The murine c-myb protein shows a striking 82% amino acid homology in the region (amino acids 71-444) where it can be compared to the published avian c-myb gene sequence. S1 nuclease protection analysis indicates extreme heterogeneity at the 5' end of steady-state murine c-myb mRNA.

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PubMed1984

Binding of insulin to its receptor impairs recognition by monoclonal anti-insulin antibodies.

The interaction between insulin and its receptor was investigated using both monoclonal and polyclonal anti-insulin antibodies. After covalent cross-linking of 125I-insulin to the insulin receptor on cultured human lymphocytes (IM-9 cells) using disuccinimidyl suberate, we inquired whether the insulin-receptor complex could be immunoprecipitated with anti-insulin antibodies. While a polyclonal guinea pig anti-insulin antiserum succeeded in immunoprecipitating receptor-bound 125I-insulin, binding to the receptor decreased the avidity of the antiserum for the insulin moiety by a factor of approximately 1000-fold. Sixteen distinct monoclonal murine anti-insulin antibodies were employed to immunoprecipitate receptor-bound 125I-insulin. Of these 16 monoclonal antibodies, only one (antibody 5.9F4) could be shown to recognize receptor-bound 125I-insulin. Moreover, even with antibody 5.9F4, binding of 125I-insulin to its receptor reduced the affinity of the antibody by a factor of 10- to 100-fold. These data strongly suggest that, when insulin binds to its receptor, the majority of the insulin molecule is unavailable for binding by anti-insulin antibodies. It seems likely that the hormone binding site on the receptor may be very large, thereby allowing for sequestration of the majority of the insulin molecule with relatively little of the hormone remaining exposed.

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PubMed1984

Two Ia.17-specific monoclonal antibodies detect the same epitope but do not share idiotype.

The clonal diversity of anti-Ia antibodies was studied by using heterologous anti-idiotypic reagents generated against two Ia.17-specific monoclonal antibodies (Mab). The Mab 10-2.16 and 10-3.6 are specific for the Ia.17 public specificity expressed on I-A molecules of the k, s, f, u, r, and j haplotypes. Competitive inhibition experiments demonstrated that 10-2.16 and 10-3.6 inhibited the binding of each other to Ia.17-positive cell targets, indicating that they detected the same or overlapping epitope(s). Identical inhibition patterns of 125I-10-2.16 binding by 10-3.6 by using Ia.17-positive target cells of four different strains indicated that inhibition was not due to steric hindrance from binding to spatially related epitopes. Rabbit anti-10-2.16 serum detected both site-specific and framework-specific 10-2.16 idiotypic determinants not shared by 10-3.6. Conversely, rabbit anti-10-3.6 serum detected 10-3.6 idiotypic determinants not shared by 10-2.16. C3H/J (I-Ak) anti-10-2.16 serum detected only 10-2.16 framework-specific idiotypes, suggesting that 10-2.16 site-specific determinants represent cellular determinants for syngeneic Ia antigens. Furthermore, anti-Ia.17 immune serum from three of 10 mice with the Igh-Cb allotype expressed the 10-2.16 framework-specific idiotype. We have demonstrated that two anti-Ia.17 Mab, 10-2.16 and 10-3.6, lack a shared idiotype, even though they detect a similar epitope. This observation and the variable expression of 10-2.16 idiotypic determinants in immune sera indicate that the B cell response to the Ia.17 epitope detected by 10-2.16 and 10-3.6 is diverse.

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PubMed1984

Anti-idiotypic regulation of an insulin-reactive T cell clone.

A series of MHC-restricted, bovine-insulin-(BI) reactive T cell clones were generated. The specificity of one group was shown to be for an insulin A-chain loop determinant; the other group apparently demonstrated specificity of a B-chain determinant and/or amino acid residue A4. Guinea pig anti-idiotypic antisera were prepared against two idiotypically related BI monoclonal antibodies of similar A-chain loop specificity. These reagents were able to modulate the antigen-specific proliferation of an insulin-reactive, A-chain loop-specific T cell clone. Because the monoclonal antibodies and the T cell clone recognize a similar molecular domain of the insulin molecule, these data suggest that the anti-idiotypic sera mimic an insulin-like determinant, perhaps by bearing an "internal image" of the antigen and thereby interfering with T cell antigen recognition. Further, these results suggest that such reagents may be useful in characterization of T cell antigen receptor specificity and lend further credence to the concept of idiotypic-anti-idiotypic regulation of the immune response.

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PubMed1983

Idiotypes on monoclonal antibodies to bovine insulin. I. Two public idiotypes on anti-bovine insulin hybridomas define idiotypically distinct families of hybridomas.

We used isoelectric focusing (IEF) to show that individual mice responding to bovine insulin (BI) present complex spectrotypes that are not conserved between individuals. Competition of 125I-BI-reactive bands with cold BI or cold pork insulin (PI), showed that the majority of the antibody response is sensitive to the two amino acid difference between BI and PI. This difference resides in the three amino acid intrachain disulfide-bonded A-loop. To begin dissection of the antibody response to BI, we prepared a panel of 30 monoclonal antibodies against insulin. Anti-idiotypic sera were produced in guinea pigs against seven of these hybridoma proteins (HP), and two, anti-5. 10C6 and anti-5.2B8, were found to define public idiotypes. The level of the 5.10C6 idiotype is controlled by genes linked to the Igh locus. Both the G.P. anti-5.10C6 and G.P. anti-5.2B8 were found to define distinct groups of idiotypically related HP. The 5.10C6 idiotype group contains three closely related members, 5.10C6, 5.5F10, and 5.4C5, which are shown to bear the same L chain by IEF. The 5.2B8 idiotype group has six members that are less closely related, defined by inhibition solid phase RIA. The two most closely related members of this family, 5.2B8 and 5.3C6, possess co-focusing L chains. Within each idiotype group, however, members were found that bind to distinct determinants on insulin. Four of the members of the 5.2B8 idiotype group appear to bind overlapping determinants on insulin that define a topographic region that includes the three amino acid A-loop. Two other HP that have been shown to bind the A-loop do not bear determinants that cross-react with the G.P. anti-5.2B8 antiserum.

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PubMedDate not supplied

A novel system to identify Myb target promoters in friend murine erythroleukemia cells.

Friend murine erythroleukemia (MEL) cells provide an early erythroid precursor model that can be induced to terminally differentiate in cell culture and has been used to study erythroid differentiation as well as multistage tumorigenesis. During the chemically induced differentiation of MEL cells, expression of the c-myb protooncogene is downregulated in a biphasic fashion and forced expression of c-myb is able to block the differentiation process, suggesting that c-myb activity may be limiting for differentiation in MEL cells. We have recently produced stable transfectants in the C19 MEL cell line that carry a dominant interfering myb allele (MEnT) under the control of an inducible mouse metallothionein I (MTH) promoter. Upon inducing expression of MEnT, transfected cells enter a differentiation program and begin to produce alpha-globin mRNA, assemble hemoglobin, and stop proliferating. Differential display was used to compare mRNA expression between parental C19 MEL cells induced to differentiate with hexamethylene bisacetamide (HMBA) and stable transfectants induced to differentiate via expression of MEnT to identify potential Myb target promoters. We identified six candidate cDNAs in this fashion and present evidence that two of these represent genes that are dependent on c-Myb activity for maximal expression in MEL cells.

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PubMedDate not supplied

Expression of a transfected human c-myc oncogene inhibits differentiation of a mouse erythroleukaemia cell line.

The Friend-virus-derived mouse erythroleukaemia (MEL) cell lines represent transformed early erythroid precursors that can be induced to differentiate into more mature erythroid cells by a variety of agents including dimethyl sulphoxide (DMSO). There is a latent period of 12 hours after inducer is added, when 80-90% of the cells become irreversibly committed to the differentiation programme, undergoing several rounds of cell division before permanently ceasing to replicate. After DMSO induction, a biphasic decline in steady-state levels of c-myc and c-myb messenger RNAs occurs. Following the initial decrease in c-myc mRNA expression, the subsequent increase occurs in, and is restricted to, the G1 phase of the cell cycle. We sought to determine whether the down-regulation is a necessary step in chemically induced differentiation. Experiments reported here indicate that expression in MEL cells of a transfected human c-myc gene inhibits the terminal differentiation process.

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