The bcl-2 family member, Bcl2a1, maps to mouse chromosome 9 and human chromosome 15.
Explore the source record for details and available documents.
Animals↗
Biomedical subjects
M B Prystowsky
Publications and source records attributed to M B Prystowsky.
Lymphocyte-specific inducible expression of potassium channel beta subunits.
Many studies have shown that voltage-gated potassium (Kv) channel activity is essential for T-lymphocyte proliferation. The IL-2-inducible neuroimmune gene, I2rf5 is the mouse homologue of the rat Kv beta 2 subunit. In this study we show that in addition to constitutive expression in adult murine brain, expression of Kv channel subunits beta 1.1 and beta 2.1 is inducible in a cloned T-helper cell line stimulated with IL-2 and in normal murine splenocytes stimulated with Con A or LPS. This expression pattern appears to be lymphocyte specific, because stimulated fibroblasts and vascular smooth muscle cells do not express Kv beta channel subunit mRNA. These observations suggest that Kv beta subunit expression is tissue specific and inducible in stimulated lymphocytes. Because Kv beta subunits modulate K+ channel activity, the inducible and variable expression of these subunits in lymphocytes may represent an additional regulatory mechanism for lymphocyte proliferation.
3T3 Cells↗
Murine leukemia virus infects early bone marrow progenitors in immunocompetent mice.
Chronic murine leukemia viruses (MuLVs) are retroviruses which induce leukemias/lymphomas after long latency periods. The induction of leukemia by MuLVs is complex, requiring multiple steps beginning with infection of an appropriate target cell. A number of investigators have proposed a bone marrow-thymus axis in the development of retrovirus induced T-cell lymphoma in which cells are initially infected in the bone marrow. These bone marrow cells or their progeny migrate to the thymus during the disease process. In our system using adult, immunocompetent BALB.K mice infected with E-55(+) MuLV, a similar pattern is seen; integrated virus is initially detectable in the bone marrow and spleen and only later in the thymus. In order to better understand the leukemic process, we analyzed the bone marrow from adult, immunocompetent BALB.K mice infected with the E-55(+) MuLV in bone marrow colony assays. The results from these assays demonstrate that either a pluripotent progenitor cell or an early progenitor cell is a target in the bone marrow for the virus.
Animals↗
A1, a Bcl-2 family member, prolongs cell survival and permits myeloid differentiation.
A1, a bcl-2 family member, has been identified as a hematopoietic-specific, early inducible gene. In this study it is shown that stable transfection of A1 into an interleukin-3 (IL-3)-dependent myeloid precursor cell line, 32D c13, leads to a retardation of IL-3 withdrawal-induced cell death similar to that observed with transfection of bcl-2. However, unlike bcl-2. A1 expression permits the accumulation of differentiated myeloid cells both before and after IL-3 withdrawal. Total cell accumulation, on the other hand, is considerably greater after IL-3 deprivation in the bcl-2 transfectant than in A1-expressing cells. Cells cotransfected with the two genes behave similarly to cells singly transfected with bcl-2, except that viability following IL-3 withdrawal is somewhat further enhanced. These results suggest that these two proteins have distinct roles that may be related to the divergent regulation of their expression during myeloid differentiation.
Animals↗
Identification of an essential cis-element near the transcription start site for transcriptional activation of the proliferating cell nuclear antigen gene.
Interleukin 2 (IL-2) stimulates T lymphocyte proliferation and induces the expression of proliferating cell nuclear antigen (PCNA), a processivity factor for DNA polymerase delta. Previously, deletion analysis suggested cis-element(s) in the proximal region of the PCNA promoter (-40 to +143) are required for IL-2 induction in cloned T lymphocytes. The sequence 5'-TTGCGGGC-3' located at +10 to +17 is similar to the E2F consensus binding site and is required for optimal PCNA promoter activity. In IL-2-stimulated T cells, nuclear proteins are induced to bind to this sequence as demonstrated using electrophoretic mobility shift assay (EMSA), competition EMSA, and methylation interference analysis. A 180-kDa polypeptide was detected by UV cross-linking to bind specifically to the PCNA E2F-like sequence. Our data indicate that the protein bound to the PCNA E2F-like site is not one of the transcription factor E2F proteins. Our results demonstrate that the E2F-like sequence and the protein(s) binding to it are required for optimal PCNA promoter activity and IL-2 induction of PCNA expression.
Animals↗
Isolation and characterization of BART-1: A novel balloon angioplasty responsive transcript in rat carotid arteries.
In previous studies, differential display analysis of balloon angioplasty-damaged rat carotid arteries identified a temporally expressed partial cDNA termed RC9. This message is undetectable in undamaged vessels, reaches maximal levels 3 days post-procedure, and reduces to half-maximal expression by 14 days post angioplasty. Using RC9 to screen a cDNA library, we now report the isolation and characterization of a full-length clone, termed BART-1. BART-1 is 98% homologous to RC9 and shows the same mRNA expression pattern as RC9 in rat carotid arteries subject to balloon angioplasty. Northern analysis of various rat tissues reveals tissue specificity and possible differential processing. Neither the nucleic acid nor amino acid sequences demonstrate similarity to previously reported expressed sequences. Predicted amino acid analysis reveals two strongly hydrophilic and one hydrophobic region, suggesting a type II integral membrane protein. The cDNA sequence hybridized to genomic DNA from a variety of species, suggesting evolutionary conservation. Thus, BART-1 mRNA appears to represent an inducible, tissue-specific transcript encoding a putative integral membrane protein transiently expressed in response to vascular trauma.
Amino Acid Sequence↗
Effect of non-H-2-linked genes on anti-virus immune responses and long-term survival in mice persistently infected with E-55+ murine leukemia virus.
We have previously demonstrated that BALB/c-H-2k (BALB.K) mice are susceptible to the development of thymic lymphoma induced by E-55+ murine leukemia virus (MuLV). In the present studies, C57BL/10-H-2k (B10.BR) mice were found to be resistant to E-55+ MuLV-induced lymphoma despite the fact that these mice become persistently infected. This resistance to lymphomagensis is mediated by the anti-virus immune response since immunosuppressed mice progress to develop disease. The protective immune response in B10.BR mice is bimodal with respect to time after virus infection. The early immune response results in a dramatic decrease in the number of virus-infected cells within 4-8 weeks after infection. This decrease in virus-infected cells occurs in immunocompetent mice from strains that are either resistant (B10.BR) or susceptible (BALB.K) to E-55+ MuLV-induced disease. Subsequently, susceptible mice develop an increase in infected cells, whereas no increase in infected cells occurs in resistant mice despite the fact that they are persistently infected. This later phase of resistance in B10.BR appears to be mediated by T cells. Since B10.BR and BALB.K both express the H-2k haplotype, resistance appears to be mediated by a non-H-2-linked gene(s). (BALB.K x B10.BR)F1 mice are resistant to disease development, indicating resistance is a dominant trait.
Animals↗
Rapamycin selectively blocks interleukin-2-induced proliferating cell nuclear antigen gene expression in T lymphocyte. Evidence for inhibition of CREB/ATF binding activities.
The macrolide rapamycin arrests T lymphocytes stimulated by interleukin-2 (IL-2) at G1/S. We have recently found that IL-2 induced an increase in the binding of discrete transcription factors of the ATF/cAMP-responsive element binding factor (CREB) family at G1/S, and that this effect was inhibited by rapamycin (Feuerstein, N., Huang, D., Hinrichs, S. H., Orten, D. J., Aiyar, N., and Prystowsky, M. B. (1995) J. Immunol. 154, 68-79). We now show, by using high resolution two-dimensional gel electrophoresis, that rapamycin inhibited selectively the synthesis of three discrete IL-2-induced soluble proteins (35 kDa/pI approximately 5, 68 kDa/pI approximately 4, 110 kDa/pI approximately 4.3). Analysis of nuclear proteins demonstrated that rapamycin selectively blocked the expression of proliferating cell nuclear antigen (PCNA), an obligate cofactor of DNA polymerase-delta, an important component for DNA replication. Rapamycin inhibited the IL-2-induced PCNA mRNA, and the murine PCNA promoter activity in IL-2-stimulated cells. Inducible CRE-binding proteins were shown previously to be required for PCNA promoter activity in IL-2-stimulated T lymphocytes. Using DNA binding gel mobility shift assay we demonstrated that rapamycin potently inhibited the binding of CREB/ATF transcription factors to CRE elements in the murine proximal PCNA promoter. These results suggest that PCNA is a preferred target in a rapamycin-sensitive transduction pathway, and that the mechanism by which rampamycin inhibits PCNA gene expression may involve the inhibition of the interaction of CREB/ATF transcription factors with CRE elements in the proximal PCNA promoter.
Activating Transcription Factors↗
Genomic organization and genetic mapping of the neuroimmune gene I2rf5 to mouse chromosome 4.
The nervous and immune systems share many functional and molecular similarities, including shared surface antigens, secretions of soluble factors, and cross-modulatory effects. We have identified previously a novel mRNA termed F5, which is expressed only in activated T lymphocytes and mature, postmitotic neurons. Tissue specificity and sequence conservation suggest an important function for F5 in T-lymphocyte proliferation and neuronal maturation. The F5 gene product is an evolutionarily conserved, cytoskeletal-associated phosphoprotein. A full-length mouse genomic clone has been isolated. The protein coding region of the F5 gene is approximately 16 kb in length and is composed of 13 coding exons. The gene encoding F5, termed I2rf5, was mapped using interspecies mouse crosses in close proximity to a number of genes associated with neuronal defects on distal chromosome 4.
Animals↗
Regulation of cAMP-responsive enhancer binding proteins during cell cycle progression in T lymphocytes stimulated by IL-2.
IL-2 stimulates the proliferative response of various lymphoid cells. Previous studies showed an increase in intracellular levels of cAMP concomitant with an increase in phosphorylation of discrete proteins by protein kinase A at late G1 phase in mitogen-stimulated lymphocytes. Thus, experiments were undertaken to study nuclear proteins that bind to the cAMP-responsive enhancer (CRE) in cloned T lymphocytes stimulated with IL-2. With the use of a 32P-labeled CRE consensus sequence in a DNA binding gel mobility shift assay, we showed that IL-2 stimulation resulted in the induction of two major DNA-protein complexes at late G1/S during the cell cycle. This binding was competed in a dose-dependent manner by a nonlabeled CRE oligonucleotide but was not competed by a nonlabeled AP-1 oligonucleotide. Rapamycin, a potent immunosuppressant, which arrests IL-2-stimulated T lymphocytes at G1/S, inhibited the IL-2-induced CRE binding activities concomitantly with inhibition of DNA synthesis. By using specific Abs in a gel mobility shift assay, we identified two known CREB/ATF transcription factors in the IL-2-induced CRE complexes: the CRE binding factor (CREB), and ATF1. The induction of CREB binding by IL-2 was not associated with an increase in its abundance but was associated with a major increase in CREB phosphorylation that was particularly prominent at late G1/S. However, we found that G1/S progression induced by IL-2 was not associated with an increase in the intracellular levels of cAMP. These results suggest that 1) the transcription factors CREB and ATF1 and possibly other CRE binding proteins may have an important role in the modulation of specific gene expression at G1/S during cell cycle progression induced by IL-2. 2) The involvement of these CRE binding transcription factors in IL-2-stimulated cells is regulated via a mechanism that is not cAMP dependent.
Activating Transcription Factor 1↗
CCAAT-box contributions to human thymidine kinase mRNA expression.
In order to examine the role of two inverted CCAAT boxes near the start of transcription of the human thymidine kinase (TK) gene, a series of constructs were prepared in which one or both CCAAT boxes were deleted or mutated. These altered promoters (1.2 kb of 5'-flanking sequence) were used to express a TK minigene containing the first two exons and introns followed by the remainder of the cDNA. RNA blots were prepared from stable cell lines of ts13 cells containing these constructs under three conditions: 1) serum deprived cells, 2) serum stimulated cells, and 3) cells that had been stimulated with serum, but were arrested in the G1 phase of the cell cycle by the temperature sensitive mutation carried by these cells. TK mRNA expression from each construct was suppressed by the temperature sensitive block to cell cycle progression. Measurement of protein expression from the various altered TK promoters indicated that both CCAAT boxes contribute to promoter strength. These experiments also suggested that the two CCAAT boxes were not equivalent and that the distal CCAAT could substitute for the proximal CCAAT, but the converse was not true.
Animals↗
Protein that binds to the distal, but not to the proximal, CCAAT of the human thymidine kinase gene promoter.
Mobility shift assays were used to examine protein binding to the human TK gene CCAAT boxes. Similar protein binding patterns were observed with probes containing either the proximal or distal CCAAT. However, probes containing both CCAAT boxes in which one of the CCAAT boxes was inactivated by mutation did not demonstrate identical binding patterns. One of the complexes formed with the longer probes was only observed when the distal CCAAT was intact. This species was not formed with probes that only contained an intact proximal CCAAT, and its formation could only be competed by oligonucleotides containing the distal CCAAT motif. This observation reveals the existence of a protein that can bind to the distal, but not to the proximal, CCAAT of the human TK promoter. This protein may account for the previous observation that the two CCAAT motifs are not functionally equivalent. The protein that binds to the distal, but not to the proximal, CCAAT (DTK-CBP) was also present in two human cell lines. Significantly more DTK-CBP was present in nuclear extracts of HepG2 and WI38 cells than in TK-ts13 cells. However, this protein was not observed in three different murine cell lines and one primary culture. Its abundance in some human cell lines suggests it might modulate the expression of human TK mRNA in cells that express this protein.
3T3 Cells↗
Environmental lighting alters the infection process in an animal model of AIDS.
In this study, we examined the effects of altered environmental lighting on the infection process of a murine leukemia virus, E-55(+), which induces a thymic lymphoma/leukemia in 100% of BALB.K mice inoculated as adults. One to two weeks after inoculation, high levels of proviral DNA are usually found. This is followed by an asymptomatic period of many weeks during which proviral DNA becomes essentially undetectable. Leukemia develops approximately 28 weeks postinoculation. In this experiment, one group of mice was exposed a consistent 10L: 14D cycle while a second was maintained in constant light (LL). A third group was exposed to a rotating cycle characterized by phase shifting a 10L: 14D cycle every three 24-h days (rLD). All cycles began 2 weeks prior to inoculation and were maintained thereafter. Animals were sacrificed at 1, 5, 10, and 15 weeks, and hematopoietic tissue was examined for proviral DNA content. At 1 week, LL- and rLD-exposed animals showed considerably less proviral DNA in bone marrow and spleen compared with controls. At 15 weeks, thymuses from controls were showing signs of infection whereas tissue from LL and rLD mice remained at background levels. We conclude that environmental lighting does alter the infective pattern displayed by this retrovirus, although whether this effect is mediated by changes in the target stem cells or through immunoenhancement has not yet been determined.
Acquired Immunodeficiency Syndrome↗
Differential effects of monoclonal antibodies on activating transcription factor-1 and cAMP response element binding protein interactions with DNA.
Activating transcription factor-1 (ATF1) and cAMP response element binding protein (CREB) have been implicated in cAMP-, calcium-, and virus-induced transcriptional alterations. Although CREB and ATF1 share extensive homology, they appear to mediate distinct cellular functions. We investigated the effect on DNA binding and in vitro transcription of four monoclonal antibodies (mAb) that bound to domains in either the regulatory region (mAb 1 and 3) or unique regions near the DNA-binding domains (mAb 4 and 5) of ATF1.mAb 1 and 3 supershifted both ATF1 and CREB in a DNA binding assay but did not affect in vitro transcription. mAb 4 prevented ATF1-DNA binding while supershifting CREB.DNA complexes and inhibited in vitro transcription by 95% from the CRE-containing murine proliferating cell nuclear antigen promoter. mAb 5 reacted specifically with ATF1 and did not prevent DNA binding or affect in vitro transcription. The mAb 4 epitope was located within ATF1 amino acid residues 205-219, including the first 3 basic residues in the putative DNA-binding domain. Secondary structural analysis predicted that this region comprises a transition site from alpha-helix to a turn-like conformation in ATF1. The transition to turn-like motifs is predicted to occur in CREB after 5 additional residues, with a correspondingly longer alpha-helical domain. Although regulatory domains distinct from DNA binding regions are thought to account for most of the differences in activity of members of the CREB subfamily, our results suggest that small structural variations adjacent to DNA binding regions may also contribute to the distinct functional activities of ATF1 and CREB.
Activating Transcription Factor 1↗
Selective induction of the beta chemokine C10 by IL-4 in mouse macrophages.
The beta chemokines are a family of 8- to 12-kDa leukocyte chemoattractants that are typically produced by activated macrophages or lymphocytes. We examined the expression in primary macrophages of a recently described, and as yet functionally uncharacterized, murine beta chemokine, C10, and contrasted its regulation with that of several other beta chemokines. Although three other beta chemokines, macrophage inflammatory protein-1 alpha (MIP-1 alpha), JE, and RANTES, were all induced by LPS treatment of bone marrow-derived macrophages (BMM) and/or resident peritoneal macrophages (RPM), LPS stimulation of C10 was never observed. Conversely, IL-3 and granulocyte macrophage-CSF (GM-CSF) strongly induced C10 in both macrophage populations, whereas MIP-1 alpha and RANTES showed a weaker induction restricted to BMM. JE was strongly induced but only in BMM. Finally, IL-4 strongly induced C10 in a dose-dependent manner in both BMM and RPM but failed to stimulate any of the other three beta chemokines. The accumulation of C10 protein in culture supernatants paralleled the induction of mRNA, and the combination of IL-4 and GM-CSF led to enhanced protein levels. The expression of the C10 message in response to cytokines was completely blocked by cycloheximide, whereas the other three chemokines were all overexpressed in the presence of this inhibitor. These results demonstrate a sharp divergence between the regulation of C10 expression and that of other chemokines and suggest that this molecule may have distinct functions in host defense.
Amino Acid Sequence↗
Prolactin as a second messenger for interleukin 2.
The generation of an effective immune response requires the selection and proliferation of antigen-specific lymphocytes. Activation of at least some T lymphocytes involves the sequential stimulation of cell surface receptors, including the T-cell receptor for antigen, the interleukin 2 receptor, and the prolactin receptor. Prolactin signaling in T lymphocytes appears to utilize a classical receptor-mediated kinase cascade and a novel peptide hormone activation pathway involving nuclear translocation.
Antigens↗
Promoter activity of the proliferating-cell nuclear antigen gene is associated with inducible CRE-binding proteins in interleukin 2-stimulated T lymphocytes.
The proliferating-cell nuclear antigen (PCNA) gene encodes an auxiliary factor of DNA polymerase delta and functions in DNA replication during S phase. It is expressed at much higher levels in proliferating cells than in quiescent cells. We have studied the regulatory role of the 5'-flanking sequence of the murine PCNA gene in interleukin 2 (IL-2)-responsive cloned T cells (L2). Analysis of a set of deletion constructs in transient transfection assays measuring heterologous reporter gene (luciferase) activity demonstrated that the 182-bp 5'-flanking region provides full promoter activity in IL-2-stimulated L2 cells. While many elements contribute to PCNA promoter strength in IL-2-stimulated cells, the largest decrease in activity occurred with deletion of the tandem CRE (cyclic AMP response element) binding sites located at nucleotides -37 to -52. With a gel mobility shift assay, several IL-2-inducible DNA-protein complexes were detected, including CREB (CRE-binding) and ATF1 (activating transcription factor) proteins that are specific for the PCNA-CRE sequence. Methylation interference analysis confirmed specific binding of these proteins to the CRE sites. Mutation at the PCNA-CRE motif abolishes IL-2-inducible binding and reduces substantially PCNA promoter activity. These results indicate that IL-2-stimulated PCNA transcription may be partially mediated by these CRE-binding proteins.
Animals↗
Characterization of A1, a novel hemopoietic-specific early-response gene with sequence similarity to bcl-2.
Granulocyte-macrophage colony-stimulating factor (GM-CSF) stimulates hemopoietic cell proliferation, differentiation, and functional activation by inducing the expression of specific genes. As part of an investigation of the regulation of gene expression by GM-CSF, we have previously identified a novel murine GM-CSF-inducible gene, A1. In this report, we present the complete nucleotide sequence of the A1 mRNA as well as a portion of the 5' flanking region, and describe the expression pattern of the gene. The results demonstrate that A1 is a hemopoietic tissue-specific gene that is expressed in several hemopoietic cell lineages, including T-helper lymphocytes, macrophages, and neutrophils. In murine bone marrow-derived macrophages, A1 gene expression is rapidly and transiently induced by GM-CSF, and the induction was independent of de novo protein synthesis. In addition to GM-CSF, a transient induction of A1 mRNA accumulation was observed in response to LPS in macrophages. This induction is not mediated by IL-1 alpha or IL-6, neither of which stimulate A1. In the myeloid precursor cell line, 32D cl3, A1 gene expression is stably induced during granulocyte colony-stimulating factor-stimulated myeloid cell differentiation. The A1 message encodes a predicted polypeptide with an M(r) of 20,024 and no signal peptide. The peptide sequence contains a region of 80 amino acids that shows similarity to bcl-2 and to the recently described bcl-2-related gene, MCL1. These data demonstrate that A1 is a novel early-response gene whose expression is associated with a variety of stimuli and occurs in several hemopoietic cell types.
Amino Acid Sequence↗