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Published on: December 9, 2016
Identification of genes leading to glucocorticoid-induced leukemic cell death
E B Thompson1, M S Webb, A L Miller
1The University of Texas Medical Branch, Department of Human Biological Chemistry & Genetics, Galveston, Texas 77555-1068, USA. bthompso@utmb.edu
Abstract:
Glucocorticoidal steroids (GC) are capable of causing apoptotic death of many varieties of lymphoid cells; consequently, GC are used in therapy for many lymphoid malignancies. Gene transcription in the GC-treated cells is required for subsequent apoptosis, but only a few of the actual genes involved have been identified. We employed gene microarray analysis to find the network of genes involved in GC-evoked cell death, using three clones derived from the CEM lymphoid leukemia cell line. Clone C1-15 was resistant to GC-evoked apoptosis, although not necessarily to GC-induced gene transcription; the other two underwent apoptosis in the presence of GC. Clone C7-14 was subcloned from the apoptosis-sensitive parental C7 clone to establish karyotypic uniformity. The second sensitive clone, C1-6, was a spontaneous revertant from parental resistant clone C1. A period of > or = 24 h in the constant presence of receptor-occupying concentrations of synthetic GC dexamethasone (Dex) was necessary for apoptosis to begin. To identify the steps leading to this dramatic event, we identified the changes in gene expression in the 20-h period preceding the onset of overt apoptosis. Cells in the log phase of growth were treated with 10(-6) M Dex, and 2-20 h later, mRNA was prepared and analyzed using the Affymetrix HG_U95Av2 chip, containing probes for about 12,600 genes. Of these, approximately 6,000 were expressed above background. Comparisons of the basal and expressed genes in the three clones led to several conclusions: The Dex-sensitive clones shared the regulation of a limited set of genes. The apoptosis-resistant clone C1-15 showed Dex effects on a largely different set of genes. Promoter analysis of the regulated genes suggested that primary gene targets for GC often lack a classic GC response element.
Insights
Glucocorticoids induce apoptosis in lymphoid cells, but the specific genes involved are unclear. This study used gene microarrays to identify genes regulated by glucocorticoids in sensitive and resistant lymphoid leukemia cell lines.
Area of Science:
- Molecular Biology
- Cell Biology
- Genomics
Background:
- Glucocorticoids (GC) induce apoptosis in lymphoid cells, a mechanism utilized in treating lymphoid malignancies.
- The specific genes and transcriptional networks mediating GC-induced apoptosis remain largely unidentified.
Purpose of the Study:
- To identify the gene expression network involved in glucocorticoid-induced apoptosis in lymphoid leukemia cells.
- To compare gene expression patterns between GC-sensitive and GC-resistant cell lines.
Main Methods:
- Gene microarray analysis (Affymetrix HG_U95Av2 chip) of three CEM lymphoid leukemia cell line clones (two sensitive, one resistant) treated with dexamethasone (Dex).
- Analysis of mRNA expression changes in the 20-hour period preceding overt apoptosis.
- Promoter analysis of regulated genes.
Main Results:
- Dex-sensitive clones exhibited coordinated regulation of a distinct set of genes compared to the resistant clone.
- The apoptosis-resistant clone displayed Dex-induced gene expression changes in a largely different set of genes.
- Primary GC target genes frequently lacked a classic GC response element.
Conclusions:
- A specific network of genes is regulated during GC-induced apoptosis in lymphoid cells.
- Differences in gene regulation correlate with GC sensitivity/resistance.
- GC may target genes lacking canonical response elements, suggesting novel regulatory mechanisms.
