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- Creator:
- Beach, Jessica Anne, Hovanessian, Rebeka, Nary, Laura J., and Medh, Rheem D.
- Description:
- In Caenorhabditiselegans, motorneuron apoptosis is regulated via a ces-2–ces-1–egl-1 pathway. We tested whether human CEM lymphoblastic leukemia cells undergo apoptosis via an analogous pathway. We have previously shown that E4BP4, a ces-2 ortholog, mediates glucocorticoid (GC)-dependent upregulation of BIM, an egl-1 ortholog, in GC-sensitive CEM C7-14 cells and in CEM C1-15mE#3 cells, which are sensitized to GCs by ectopic expression of E4BP4. In the present study, we demonstrate that the human ces-1 orthologs, SLUG and SNAIL, are not significantly repressed in correlation with E4BP4 expression. Expression of E4BP4 homologs, the PAR family genes, especially HLF, encoding a known anti-apoptotic factor, was inverse to that of E4BP4 and BIM. Expression of pro- and anti-apoptotic genes in CEM cells was analyzed via an apoptosis PCR Array. We identified BIRC3 and BIM as genes whose expression paralleled that of E4BP4, while FASLG, TRAF4, BCL2A1, BCL2L1, BCL2L2 and CD40LG as genes whose expression was opposite to that of E4BP4.
- Resource Type:
- Article
- Identifier:
- 0006-291X
- Campus Tesim:
- Northridge
- Creator:
- Medh, Rheem D.
- Description:
- Recent advances in gene microarray technology have facilitated global analyses of gene expression profiles in normal and malignant immune cells. Great strides have been made in our understanding of molecular differences among various types of immune cells, the process of T and B cell activation, and the genomic changes that convert normal cells to malignant ones. Genomic analysis has become a crucial aspect of cancer classification, diagnosis, therapy, and prognosis. This technology has the potential to reveal the comprehensive transcriptional alterations that dictate fundamental biological processes such as signal transduction in response to specific stimuli, cell growth, differentiation, and apoptosis. While reaping the benefits of genomic analyses, it is important to realize its limitations with respect to accuracy of interpretation, reproducibility, and signal detection. It is crucial to optimize signals for individual probe-target pairs and to develop a uniform set of criteria for data analyses. The development of a public-access database of results from individual laboratories will pave the way for identifying discrepancies and advancing scientific breakthroughs.
- Resource Type:
- Article
- Identifier:
- 0163-769X
- Campus Tesim:
- Northridge
- Creator:
- Saenz, G. Jonatan, Medh, Rheem D., Gisis, Andrew D., and Hovanessian, Rebeka
- Description:
- Glucocorticoids (GCs) are known to induce apoptosis of leukemia cells via gene regulatory changes affecting key pro-and anti-apoptotic genes. Three genes previously implicated in GC-evoked apoptosis in the CEM human T-cell leukemia model, RCAN1, E4BP4 and BIM, were studied in a panel of human lymphoid and myeloid leukemia cell lines. Of the two RCAN1 transcripts, the synthetic GC Dexamethasone (Dex) selectively upregulates RCAN1-1, but not RCAN1-4, in GC-susceptible Sup-B15, RS4;11, Kasumi-1 cells but not in GC-resistant Sup T1 and Loucy cells. E4BP4 and BIM regulation correlated with that of RCAN1-1. A putative GRE and four EBPREs were identified within 15bp upstream from the transcription start site of RCAN1-1. GC-refractory CEM C1-15 cells sensitized to GC-evoked apoptosis by ectopic E4BP4 expression, CEM C1-15mE#3, showed restored RCAN1-1 upregulation, suggesting that RCAN1-1 is a downstream target of E4BP4. A model for coordinated regulation of RCAN1-1, E4BP4 and BIM, and their role in GC-evoked apoptosis is proposed.
- Resource Type:
- Article
- Identifier:
- 0006-291X
- Campus Tesim:
- Northridge
- Creator:
- Yarborough, Jonathan R., Medh, Rheem D., Shankar, Deepa B., Todd, Jennifer, Lee, Lee H., Okafor, Chiedozie, Morris, Devin, Kirzner, Jonathan D., Sakamoto, Kathleen M., Perez, Winder, Chu, Tin-Chun, Murray, Sean R., Nary, Laura J., and Priceman, Saul J.
- Description:
- Glucocorticoid (GC)-evoked apoptosis of T-lymphoid cells is preceded by increases in the intracellular Ca2+ concentration ([Ca2+]i), which may contribute to apoptosis. This report demonstrates that GC-mediated upregulation of the bZIP transcriptional repressor gene, E4BP4, is dependent on [Ca2+]i levels, and correlates with GC-evoked apoptosis of GC-sensitive CEM-C7-14 cells. Calcium chelators EGTA and BAPTA reduced [Ca2+]i levels and protected CEM-C7-14 cells from Dex-evoked E4BP4 upregulation as well as apoptosis. In the GC-resistant sister clone, CEM-C1-15, Dex treatment did not induce [Ca2+]i levels, E4BP4 expression or apoptosis, however, the calcium ionophore A23187 restored Dex-evoked E4BP4 upregulation and apoptosis. CEM-C7-14 cells were more sensitive to GC-independent increases in [Ca2+]i levels by thapsigargin, and a corresponding increase in E4BP4 expression and cell death, compared to CEM-C1-15 cells, suggesting a direct correlation between [Ca2+]i levels, E4BP4 expression, and apoptosis.
- Resource Type:
- Article
- Identifier:
- 0006-291X
- Campus Tesim:
- Northridge
- Creator:
- Medh, Rheem D., Holland, Eli, Hirakawa, Yasuko, Hovanessian, Rebeka, and Beach, Jessica Anne
- Description:
- BACKGROUND: Synthetic GCs serve as therapeutic agents for some lymphoid leukemias because of their ability to induce transcriptional changes via the GC receptor (GR) and trigger apoptosis. Upregulation of the BH3-only member of Bcl-2 family proteins, Bim, has been shown to be essential for GC-evoked apoptosis of leukemic lymphoblasts. Using human T cell leukemic sister clones CEM-C7-14 and CEM-C1-15, we have previously shown that the bZIP transcriptional repressor, E4BP4, is preferentially upregulated by GCs in CEM-C7-14 cells that are susceptible to GC-evoked apoptosis, but not in refractory CEM-C1-15 cells. E4BP4 is an evolutionarily conserved member of the PAR family of bZIP transcription factors related to the C. elegans death specification gene ces2. RESULTS: Mouse E4BP4 was ectopically expressed in CEM-C1-15 cells, resulting in sensitization to GC-evoked apoptosis in correlation with restoration of E4BP4 and Bim upregulation. shRNA mediated modest knockdown of E4BP4 in CEM-C7-14 cells resulted in concomitant reduction in Bim expression, although GC-evoked fold-induction and sensitivity to apoptosis was similar to parental cells. CONCLUSION: Data presented here suggest that GC-mediated upregulation of E4BP4 facilitates Bim upregulation and apoptosis of CEM cells. Since the Bim promoter does not contain any consensus GRE or EBPRE sequences, induction of Bim may be a secondary response.
- Resource Type:
- Article
- Identifier:
- 1750-2187
- Campus Tesim:
- Northridge
- Creator:
- Saumell, Maria, Medh, Rheem D., Shankar, Deepa B., Fernandez, Silvia, Morris, Devin, Abrego, Bernardo, Hurtado, Ferran, Sakamoto, Kathleen M., Priceman, Saul J., Fabila-Monroy, Ruy, Flores-Penaloza, David, Sacristan, Vera, Kirzner, Jonathan D., and Nary, Laura J.
- Description:
- Let P be a set of n points in the plane. A geometric proximity graph on P is a graph where two points are connected by a straight-line segment if they satisfy some prescribed proximity rule. We consider four classes of higher order proximity graphs, namely, the k-nearest neighbor graph, the k-relative neighborhood graph, the k-Gabriel graph and the k -Delaunay graph. For k=k= (k=1k=1 in the case of the k-nearest neighbor graph) these graphs are plane, but for higher values of k in general they contain crossings. In this paper, we provide lower and upper bounds on their minimum and maximum number of crossings. We give general bounds and we also study particular cases that are especially interesting from the viewpoint of applications. These cases include the 1-Delaunay graph and the k-nearest neighbor graph for small values of k.
- Resource Type:
- Article
- Identifier:
- 0925-7721
- Campus Tesim:
- Northridge
- Creator:
- Johnson, Betty H, Medh, Rheem D., Thompson, E.Brad, Fofanov, Yuriy, Miller, Aaron L., Luxon, Bruce A, Webb, M.Scott, Li, Tongbin, and Wood, Thomas G
- Description:
- Three closely related clones of leukemic lymphoid CEM cells were compared for their gene expression responses to the glucocorticoid dexamethasone (Dex). All three contained receptors for Dex, but only two responded by undergoing apoptosis. After a time of exposure to Dex that ended late in the interval preceding onset of apoptosis, gene microarray analyses were carried out. The results indicate that the expression of a limited, distinctive set of genes was altered in the two apoptosis-prone clones, not in the resistant clone. That clone showed altered expression of different sets of genes, suggesting that a molecular switch converted patterns of gene expression between the two phenotypes: apoptosis-prone and apoptosis-resistant. The results are consistent with the hypothesis that altered expression of a distinctive network of genes after glucocorticoid administration ultimately triggers apoptosis of leukemic lymphoid cells. The altered genes identified provide new foci for study of their role in cell death.
- Resource Type:
- Article
- Identifier:
- 0888-7543
- Campus Tesim:
- Northridge
- Creator:
- Medh, Rheem D., Shankar, Deepa B., Morris, Devin, Kirzner, Jonathan D., Sakamoto, Kathleen M., Priceman, Saul J., and Nary, Laura J.
- Description:
- Glucocorticoid (GC)-evoked apoptosis of T-lymphoid cells is preceded by increases in the intracellular Ca2+ concentration ([Ca2+]i), which may contribute to apoptosis. This report demonstrates that GC-mediated upregulation of the bZIP transcriptional repressor gene, E4BP4, is dependent on [Ca2+]i levels, and correlates with GC-evoked apoptosis of GC-sensitive CEM-C7-14 cells. Calcium chelators EGTA and BAPTA reduced [Ca2+]i levels and protected CEM-C7-14 cells from Dex-evoked E4BP4 upregulation as well as apoptosis. In the GC-resistant sister clone, CEM-C1-15, Dex treatment did not induce [Ca2+]i levels, E4BP4 expression or apoptosis, however, the calcium ionophore A23187 restored Dex-evoked E4BP4 upregulation and apoptosis. CEM-C7-14 cells were more sensitive to GC-independent increases in [Ca2+]i levels by thapsigargin, and a corresponding increase in E4BP4 expression and cell death, compared to CEM-C1-15 cells, suggesting a direct correlation between [Ca2+]i levels, E4BP4 expression, and apoptosis.
- Resource Type:
- Article
- Identifier:
- 0006-291X
- Campus Tesim:
- Northridge
- Creator:
- Metzenberg, Stan, Medh, Rheem D., and Hirakawa, Yasuko
- Description:
- BackgroundQuantitative Polymerase Chain Reaction (qPCR) is a collection of methods for estimating the number of copies of a specific DNA template in a sample, but one that is not universally accepted because it can lead to highly inaccurate (albeit precise) results. The fundamental problem is that qPCR methods use mathematical models that explicitly or implicitly apply an estimate of amplification efficiency, the error of which is compounded in the analysis to unacceptable levels.ResultsWe present a new method of qPCR analysis that is efficiency-independent and yields accurate and precise results in controlled experiments. The method depends on a computer-assisted deconvolution that finds the point of concordant amplification behavior between the "unknown" template and an admixed amplicon standard. We apply the method to demonstrate dexamethasone-induced changes in gene expression in lymphoblastic leukemia cell lines.ConclusionsThis method of qPCR analysis does not use any explicit or implicit measure of efficiency, and may therefore be immune to problems inherent in other qPCR approaches. It yields an estimate of absolute initial copy number of template, and controlled tests show it generates accurate results.
- Resource Type:
- Article
- Identifier:
- 1471-2199
- Campus Tesim:
- Northridge
- Creator:
- Medh, Rheem D., Nary, Laura J., Hovanessian, Rebeka, and Beach, Jessica Anne
- Description:
- In Caenorhabditiselegans, motorneuron apoptosis is regulated via a ces-2–ces-1–egl-1 pathway. We tested whether human CEM lymphoblastic leukemia cells undergo apoptosis via an analogous pathway. We have previously shown that E4BP4, a ces-2 ortholog, mediates glucocorticoid (GC)-dependent upregulation of BIM, an egl-1 ortholog, in GC-sensitive CEM C7-14 cells and in CEM C1-15mE#3 cells, which are sensitized to GCs by ectopic expression of E4BP4. In the present study, we demonstrate that the human ces-1 orthologs, SLUG and SNAIL, are not significantly repressed in correlation with E4BP4 expression. Expression of E4BP4 homologs, the PAR family genes, especially HLF, encoding a known anti-apoptotic factor, was inverse to that of E4BP4 and BIM. Expression of pro- and anti-apoptotic genes in CEM cells was analyzed via an apoptosis PCR Array. We identified BIRC3 and BIM as genes whose expression paralleled that of E4BP4, while FASLG, TRAF4, BCL2A1, BCL2L1, BCL2L2 and CD40LG as genes whose expression was opposite to that of E4BP4.
- Resource Type:
- Article
- Identifier:
- 0006-291X
- Campus Tesim:
- Northridge