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Biomedical subjects

D Reisman

Publications and source records attributed to D Reisman.

At least 19 recordsLinked to original sources
PubMed2006

Increased p53 transcription prior to DNA synthesis is regulated through a novel regulatory element within the p53 promoter.

p53 mRNA levels are tightly regulated during the cell cycle with its transcription being induced prior to DNA synthesis. However, the mechanism controlling this regulation is not well defined. Through characterizing an additional 1000 bp of upstream DNA sequences of the murine p53 gene, we identified new positive and negative regulatory elements. Furthermore, we found a trans-acting factor(s) that binds within a positive cis-acting element (-972/-953) in a manner indicative of regulation during the cell cycle. When Swiss3T3 cells are arrested by serum depletion p53 mRNA levels decrease and binding of this regulatory factor(s) to the promoter is reduced. Upon serum stimulation, the regulatory factor(s) binds the promoter and p53 mRNA levels increase prior to the cells entering S phase. When the factors are experimentally sequestered from the promoter or when the regulatory element is deleted from the promoter, p53 promoter activity is reduced. There is no further reduction in p53 promoter activity upon serum depletion and the kinetics of induction upon serum stimulation is delayed by approximately 5 h. These findings indicate that a factor(s) binding within the -972/-953 regulatory element on the p53 promoter is important for the proper regulation of p53 mRNA expression in response to mitogen stimulation. Our initial findings indicate that a member of the C/EBP family of transcription factors may play a role in this regulation.

3T3 Cells↗
PubMed2004

Laser-driven plasma loader for shockless compression and acceleration of samples in the solid state.

A new method for shockless compression and acceleration of solid materials is presented. A plasma reservoir pressurized by a laser-driven shock unloads across a vacuum gap and piles up against an Al sample thus providing the drive. The rear surface velocity of the Al was measured with a line VISAR, and used to infer load histories. These peaked between approximately 0.14 and 0.5 Mbar with strain rates approximately 10(6)-10(8) s(-1). Detailed simulations suggest that apart from surface layers the samples can remain close to the room temperature isentrope. The experiments, analysis, and future prospects are discussed.

Journal Article↗
PubMed2001

Effects of varying task constraints on solutions to joint coordination in a sit-to-stand task.

The question of how multijoint movement is controlled can be studied by discovering how the variance of joint trajectories is structured in relation to important task-related variables. In a previous study of the sit-to-stand task, for instance, variations of body segment postures that leave the position of the body's center of mass (CM) unchanged were significantly greater than variations of body segment posture that varied the CM position. The present experiments tested the hypothesis that such structuring of joint configuration variability is accentuated when the mechanical or perceptual task demands are made more challenging. Six subjects performed the sit-to-stand task without vision (eyes closed), either on a normal or on a narrow support surface. An additional constraint on the postural task was introduced in a third condition by requiring subjects to maintain light touch (less than 1 N) with the fingertips while coming to a standing position on the narrow base of support. The joint configurations observed at each point in normalized time were analyzed with respect to trial-to-trial variability. The task variables CM and head position were used to define goal-equivalent sets of joint configurations ("uncontrolled manifolds," UCMs) within which variation of joint configuration leaves the task variables unchanged. The variability of joint configurations across trials was decomposed into components that did not affect (within the UCM) and that did affect (orthogonal to the UCM) the values of these task variables. Our results replicate the earlier finding of much larger variability in directions of joint space that leave the CM unchanged compared with directions that affect CM position. This effect was even more pronounced here than in the previous experiment, probably because of the more difficult perceptual conditions in the current study (eyes closed). When the mechanical difficulty of the task was increased, the difference between the two types of joint variability was further accentuated, primarily through increase in goal-equivalent variance. This provides evidence for the hypothesis that under challenging task constraints increased variability is selectively directed into task-irrelevant degrees of freedom. Because differential control along different directions of joint space requires coordination among joint angles, this observation supports the view that the CNS responds to increased task difficulty through enhanced coordination among degrees of freedom. The adaptive nature of this coordination is further illustrated by the similar enhanced use of goal-equivalent joint combinations to achieve a stable CM position when subjects stood up under the additional constraint of maintaining light touch with the fingertips. This was achieved by channeling goal-equivalent variability into different directions of joint configuration space.

Adult↗
PubMed2001

Cloning and characterization of murine p53 upstream sequences reveals additional positive transcriptional regulatory elements.

Transcriptional regulation of the p53 gene plays an important role leading to elevated expression of mutant p53 alleles in tumor cells. In addition, alterations in p53 transcription levels occur in response to changes in the cell cycle. Previous work had identified a number of regulatory sites at the 5'-end of the murine p53 promoter. During the characterization of the 5'-end of the cloned murine p53 promoter, we identified a 28 bp positive regulatory element that participates in three distinct DNA-protein complexes. The binding by nuclear factors to each one of these sites contributes to the overall activity of the p53 promoter. One site is a potential recognition sequence for members of the ETS family of transcription factors, which are known regulators of the human p53 promoter. Since six nucleotides in the middle of this required element were not present in the previously published sequence of the murine promoter, we recloned this region from C57/BL6 cells and confirmed their presence in the genome. The removal of this regulatory element completely abolishes p53 promoter activity.

3T3 Cells↗
PubMed2000

Compromised HOXA5 function can limit p53 expression in human breast tumours.

Expression of the p53 gene protects cells against malignant transformation. Whereas control of p53 degradation has been a subject of intense scrutiny, little is known about the factors that regulate p53 synthesis. Here we show that p53 messenger RNA levels are low in a large proportion of breast tumours. Seeking potential regulators of p53 transcription, we found consensus HOX binding sites in the p53 promoterS. Transient transfection of Hox/HOXA5 activated the p53 promoter. Expression of HOXA5 in epithelial cancer cells expressing wild-type p53, but not in isogenic variants lacking the p53 gene, led to apoptotic cell death. Moreover, breast cancer cell lines and patient tumours display a coordinate loss of p53 and HOXA5 mRNA and protein expression. The HOXA5 promoter region was methylated in 16 out of 20 p53-negative breast tumour specimens. We conclude that loss of expression of p53 in human breast cancer may be primarily due to lack of expression of HOXA5.

Animals↗
PubMed1999

Inhibition of the putative tumor suppressor gene TIMP-3 by tumor-derived p53 mutants and wild type p53.

The p53 gene is a tumor suppressor that regulates the expression of genes required for cell cycle arrest or apoptosis. Mutations in p53 have been observed in over 60% of all human cancers. Certain classes of mutant p53 proteins maintain some of their activities or acquire novel activities and thus may contribute to the transformed phenotype. By carrying out an analysis of differential gene expression using cDNA expression arrays, we compared the expression patterns of cells expressing no p53 to isogenic lines expressing the codon 248 Arg to Trp mutant p53 allele (R248W). In this report, we show that the R248W and D281G p53 mutants, two of the more commonly occurring mutations, as well as wild type p53, repress transcription of the tissue inhibitor of metalloproteinases type 3 (TIMP-3) gene by greater than tenfold. TIMP-3 expression has been observed to be repressed in many tumors and its reduced expression is thought to contribute to tumor metastasis and invasiveness by allowing increased activity of metalloproteinases in the extracellular matrix. Since mutant forms of p53 tend to be expressed at greatly elevated levels in many human tumors, the retention of their ability to repress TIMP-3 illustrate one mechanism by which mutant forms of the p53 gene may contribute to tumorigenesis.

Actins↗
PubMed1999

Characterization of risks posed by combustor emissions.

Risk characterization is the final step of the risk assessment process as practiced in the U.S. EPA. In risk characterization, the major scientific evidence and "bottom-line" results from the other components of the risk assessment process, hazard identification, dose-response assessment, and exposure assessment, are evaluated and integrated into an overall conclusion about the risks posed by a given situation. Risk characterization is also an iterative process; the results of a specific step may require re-evaluation or additional information to finalize the risk assessment process. Risks posed by atmospheric emissions are an example of an involuntary human health risk which typically receives a great deal of public attention. Characterization of the risks posed by atmospheric emissions typically requires the use of mathematical models to evaluate: 1) the environmental fate of emitted pollutants, 2) exposures to these pollutants, and 3) human dose-response. Integration of these models results in quantitative risk estimates. The confidence in a quantitative risk estimate is examined by evaluating uncertainty and variability within individual risk assessment components. Variability arises from the true heterogeneity in characteristics within a population or an event; on the other hand, uncertainty represents lack of knowledge about the true value used in a risk estimate. U.S. EPA's 1997 Mercury Study will illustrate some aspects of the risk characterization process as well as the uncertainty and variability encountered in the risk assessment process.

Air Pollutants↗
PubMed1999

Elevated expression of ribosomal protein genes L37, RPP-1, and S2 in the presence of mutant p53.

The wild-type p53 protein is a DNA-binding transcription factor that activates genes such as p21, MDM2, GADD45, and Bax that are required for the regulation of cell cycle progression or apoptosis in response to DNA damage. Mutant forms of p53, which are transforming oncogenes and are expressed at high levels in tumor cells, generally have a reduced binding affinity for the consensus DNA sequence. Interestingly, some p53 mutants that are no longer effective at binding to the consensus DNA sequence and transactivating promoters containing this target site have acquired the ability to transform cells in culture, in part through their ability to transactivate promoters of a number of genes that are not targets of the wild-type protein. Certain p53 mutants are therefore considered to be gain-of-function mutants and appear to be promoting proliferation or transforming cells through their ability to alter the expression of novel sets of genes. Our goal is to identify genes that have altered expression in the presence of a specific mutant p53 (Arg to Trp mutation at codon 248) protein. Through examining differential gene expression in cells devoid of p53 expression and in cells that express high levels of mutant p53 protein, we have identified three ribosomal protein genes that have elevated expression in response to mutant p53. Consistent with these findings, the overexpression of a number of ribosomal protein genes in human tumors and evidence for their contribution to oncogenic transformation have been reported previously, although the mechanism leading to this overexpression has remained elusive. We show results that indicate that expression of these specific ribosomal protein genes is increased in the presence of the R248W p53 mutant, which provides a mechanism for their overexpression in human tumors.

Animals↗
PubMed1998

Transcriptional regulation of the p53 tumor suppressor gene.

Transcriptional regulation of the p53 gene plays an important role in expression of wild type p53 and in elevated expression of mutant p53 genes in tumor cells. In addition, alterations in p53 transcription levels occur in response to changes in the cell cycle. The following review describes current knowledge regarding the structures of both the human and murine p53 transcriptional regulatory regions and the factors that have been demonstrated to regulate promoter activity. The potential role of defects in transcriptional regulation of the p53 gene in oncogenic transformation is discussed.

Animals↗
PubMed1997

Anesthetic efficacy of the supplemental intraosseous injection of 3% mepivacaine in irreversible pulpitis.

OBJECTIVE: To determine the efficacy of a supplemental intraosseous injection of 3% mepivacaine in mandibular posterior teeth with irreversible pulpitis. Intraosseous injection pain, subjective heart rate increase, and pain ratings during endodontic treatment were also assessed. STUDY DESIGN: Forty-eight patients with irreversible pulpitis received conventional inferior alveolar nerve blocks. Electric pulp testing was used to determine pulpal anesthesia. Patients who were positive to the pulp testing, or negative to pulp testing but felt pain during endodontic treatment, received an intraosseous injection of 1.8 ml of 3% mepivacaine. A second intraosseous injection of 3% mepivacaine (1.8 ml) was given if the first injection was unsuccessful. RESULTS: Seventy-five percent of patients required an initial intraosseous injection because of failure to gain pulpal anesthesia. The inferior alveolar block was 25% successful; the first intraosseous injection increased success to 80%. A second intraosseous injection further increased success to 98%. These differences were significant (p < 0.05). Eight percent (4/48) of the initial intraosseous injections resulted in solution being expressed into the oral cavity: these were considered technique failures. CONCLUSIONS: For mandibular posterior teeth with irreversible pulpitis, a supplemental intraosseous injection of 3% mepivacaine increased anesthetic success. A second intraosseous injection, when necessary, further improved success.

Adolescent↗
PubMed1996

A novel transcript encoded within the 10-kb first intron of the human p53 tumor suppressor gene (D17S2179E) is induced during differentiation of myeloid leukemia cells.

Two promoters were previously shown to map to the 5'-end of the human p53 gene. p53p1 was located upstream of the first exon and is responsible for transcription of the major p53 mRNA species. p53p2 is a stronger promoter than p53p1 and was located within the 10, 738-bp first intron, approximately 1000 bp downstream of exon 1. mRNA transcripts that initiated from p53p2 were previously identified in HL-60 cells by primer extension analysis and were observed to increase in abundance during differentiation of HL-60 cells to granulocytes. By screening a cDNA library with a probe derived from sequences downstream of the p53p2 start site, we have cloned and characterized a cDNA that represents a mRNA that appears to have been initiated from the p53p2 promoter. We have designated the gene encoding this transcript Hp53int1 (the GDB designation is D17S2179E). The cDNA is 1125 bp and is polyadenylated downstream from a consensus poly(A) addition site. The entire 1125 bp is derived from intron 1 of the p53 gene, with no introns having been removed. The cDNA contains no major open reading frame although reading frame +1 contains a relatively low abundance of stop codons compared to the other two reading frames and could possibly encode a protein of 119 amino acids. Analysis of the +1 reading frame shows a region of high homology to a portion of the DNA-binding domain of NF-kappaB. These results indicate that a novel polyadenylated transcript is encoded by the first intron of the human p53 gene. Hp53int1 may be a pseudogene for a gene that may have encoded a DNA-binding protein. Alternatively, the transcript may have a function, since RNA transcripts of this gene are present in a number of human cells and their levels are induced during terminal differentiation of myeloid leukemia cells such as HL-60 and U937.

Amino Acid Sequence↗
PubMed1996

Positive and negative regulatory elements in the murine p53 promoter.

In order to understand the basis for regulated as well as de-regulated expression of the p53 tumor suppressor gene, we have focused on characterizing the transcriptional regulation of the p53 gene. Here we present evidence for the existence of two additional upstream regulatory elements in the murine p53 promoter. One of these sites maps to a region between -296 to -270 and the second one between -255 to -226 relative to the major transcription initiation site. These two sites are referred to as binding sites for PBF I and II, respectively. Nucleotide bases that have been found to be critical for the binding of nuclear factors to these sites are 5'-AGA-3' (-282 to -280) in binding site I and 5'-ACAG-3' (-246 to -243) in binding site II. Mutational analyses in conjunction with transient transfection assays indicated that the factor that binds to the region between -245 to -242 (PBF II) plays a positive regulatory role p53 promoter activity. This was demonstrated by the observation that promoter mutations that abolished binding to this site, showed a decreased level of activity as compared to the wild type promoter. In analogous experiments, mutational anlayses and transient transfection assays indicated that the factor that binds to the region between -282 to -280 (PBF I) plays a negative regulatory role in p53 promoter activity. This was demonstrated by the observation that promoter mutations that abolished binding to this site, showed an increased level of activity as compared to the wild type promoter.

3T3 Cells↗
PubMed1996

Increased rate of transcription contributes to elevated expression of the mutant p53 gene in Burkitt's lymphoma cells.

Mutations and elevated levels of the p53 tumor suppressor protein have been reported in Burkitt's lymphomas (BL). We investigated whether elevated levels of the mutant p53 protein can be due to transcriptional or posttranscriptional mechanisms. In surveying a series of B-lymphoid cell lines, we found that a high level of the p53 protein tended to reflect high steady-state levels of p53 mRNA. p53 mRNA exhibited high stability in all of the cells tested. Cycloheximide treatment of the cells showed that the low level of p53 mRNA in non-BL lymphoid cells can not be attributed to posttranscriptional regulation by a labile protein. Nuclear run-on experiments revealed that the transcription rate of the p53 gene is about four times higher in some high p53-expressing BL cell lines as compared to low p53-expressing lymphoid cell lines. Our data suggest that an increase in the rate of transcription of the p53 gene is one of the mechanisms that contribute to elevated levels of mutant p53 protein observed in many tumor cells.

Burkitt Lymphoma↗
PubMed1996

AP-1 DNA binding activity induced by hyperosmolality in the rat hypothalamic supraoptic and paraventricular nuclei.

Immediate early gene products (c-fos, c-jun and their cognates) act as transcription factors coupling physiologically relevant stimuli to long-term responses by binding to the AP-1 site in the promoter region of target genes. The induction of c-fos has been identified in the paraventricular (PVN) and supraoptic (SON) hypothalamic magnocellular nuclei after hyperosmotic stimulation by using in situ hybridization and immunocytochemistry. In this study, AP-1 DNA binding activity, an indicator of the functional form of the c-fos transcription factor, was examined in nuclear extracts prepared from these brain regions using an electrophoretic mobility shift assay and a labeled oligonucleotide containing the AP-1 consensus sequence. Two hours after hypertonic saline injection (i.p.), rats were killed and nuclear proteins were extracted from tissue punches of brain regions to assess AP-1 binding activity. Hyperosmolality induced an increase of AP-1 binding activity in nuclear protein from SON and PVN, but not striatum. This binding was competitively displaced by excess unlabeled AP-1 oligonucleotide whereas addition of increasing amounts of unlabeled SP-1 oligonucleotide (promoter site on housekeeping genes for the ubiquitous SP-1 transcription factor) did not decrease the binding. The binding protein was shown to contain c-Fos/Fra and c-Jun since addition of c-Fos/Fra antiserum formed a supershift of the DNA, protein and antibody complex, and c-Jun antibody blocked the protein DNA binding. These data suggest that hyperosmolality leads to a selective and specific increase in AP-1 DNA binding activity which may be responsible for regulating secondary target gene expression in the hypothalamic SON and PVN.

Animals↗
PubMed1995

Transcriptional repression of p53 by human T-cell leukemia virus type I Tax protein.

The human T-cell leukemia virus type I oncoprotein Tax transcriptionally deregulates a wide variety of viral and cellular genes. Tax deregulation of gene expression is mediated through interaction with a variety of structurally unrelated cellular transcription factors, as Tax does not bind DNA in a sequence-specific manner. Although most of these cellular transcription factors have been shown to mediate activation by Tax, we have recently demonstrated that members of the basic helix-loop-helix (bHLH) family of transcription factors, which play a critical role in progression through the cell cycle, mediate repression by Tax. In this report, we examined whether Tax might repress transcription of the tumor suppressor p53, as the p53 gene has recently been demonstrated to be regulated by the bHLH protein c-Myc. Furthermore, loss or inactivation of the p53 gene has been shown to be causally associated with oncogenic transformation. We show that Tax represses transcription of the p53 gene and that this repression is dependent upon the bHLH recognition element in the p53 promoter. Together, these results suggest that Tax may promote malignant transformation through repression of p53 transcription.

Cell Line↗
PubMed1995

Transforming growth factor-beta 1 inhibits cyclin D1 expression in intestinal epithelial cells.

Transforming growth factor-beta 1 (TGF-beta 1) inhibits most epithelial cell types by blocking cell cycle progression during the G1 phase. D cyclins are normally expressed during G1 and are regulators of G1 progression. One of the crucial functions of D cyclins is their ability to bind to a cyclin-dependent kinase (Cdk4). In mink lung epithelial cells, TGF-beta 1 inhibits Cdk4 expression. We have measured cell cycle progression and D cyclins and Cdk4 expression in non-transformed rat intestinal epithelial cell lines (IEC-6 and RIE-1) after TGF-beta 1 treatment. In exponentially growing cultures, TGF-beta 1 blocked DNA synthesis and suppressed cyclin D1 mRNA and protein expression, whereas the levels of cyclins D2, D3 and Cdk4 remained relatively unchanged. TGF-beta 1 was also added to G0-synchronized IEC-6 cells after serum stimulation. TGF-beta 1 prevention of G1 progression was associated with an inhibition of cyclin D1 protein expression. Cyclin D3 levels were not affected by TGF-beta 1 during G1 traverse. Our results suggest that cyclin D/Cdk4 is a crucial target of TGF-beta 1 and that regulation of this kinase is mediated through cyclin D1 in intestinal epithelial cells.

Animals↗
PubMed1995

Inducible expression of Mad accelerates growth arrest of serum deprived human glioblastoma cells.

Mad is a basic-helix-loop-helix leucine zipper protein that heterodimerizes with Max. Mad:Max heterodimers recognize the same DNA binding sites as Myc:Max heterodimers. However, Myc and Mad are thought to influence transcription and cell proliferation in opposite ways through interaction with Max. While Myc activates transcription and cell proliferation Mad represses these activities (Ayer et. al., Cell 72, 211-222, 1993). We have constructed an expression vector containing the mad cDNA cloned downstream of an inducible viral promoter. Cell lines carrying the construct were derived by transfection into human glioblastoma cells and the effect of Mad induction on their growth rate was studied. When the cells were cultured in low serum concentrations (0.1%) Mad induction resulted in accelerated growth arrest. These findings are consistent with the proposed growth repression function of the Mad protein.

Basic Helix-Loop-Helix Leucine Zipper Transcriptio↗