Project name: Homo sapiens

Description: Glucorticoids (GCs) are steroid hormones with a wide range of physiological actions in different cell types and tissues. Their ability to induce apoptosis in malignant cells of the lymphoid lineage is exploited since decades in the treatment of childhood acute lymphoblastic leukemia (ALL), the molecular mechanism of this cell death induction being however still unknown.Using an integrative approach we delineated the transcriptional response of a preB- and a T-ALL model system employing Exon microarrays and identified in vivo interactions of the transcription factor GC receptor (GR) with genes' promoters in the same samples using the ChIP-on-chip technology. We did find differences in GR-DNA interaction in the genes' promoters between the two cell systems and also in extent and kinetics of GC mediated gene regulation. The overall response was stronger in CEM-C7H2 T-ALL cells where it was also accompanied by a stronger autoup-regulation of the GR gene. Considering the differing kinetics we furthermore defined a set of common response genes. These response genes were enriched in apoptosis related gene ontology processes, and about half of them yielded also GR enrichment in the respective promoter regions.Globally, the presence of GR at gene's promoters was higher for GC induced compared to repressed genes, indicating that GR mediates gene repression by interaction with distant enhancer regions or by cross-talk with other transcription factors.In order to identify the genes' GC regulated transcript variants we analyzed the data set also at the level of individual exons and predicted first exons of GC induced isoforms. We could thus define GC regulated variants of response genes, did however also identify first exons not corresponding to any known transcript.We reconfirmed these results for the genes ATP4B, GPR98, TBCD and ZBTB16 and identified for the latter also a novel exon employing 5' nested amplification of cDNA ends. Expression and regulation of this and all downstream exons of ZBTB16 was further verified by real time RT-PCR in 4 different preB-ALL cell lines. Thus, besides potential other novel gene isoforms, GCs induce a novel short transcript variant of ZBTB16 in preB-ALL cells.Finally, the implemented bioinformatic methods may be useful also for other high density microarray data sets to identify alternative splice events and transcript variant expression.Overall design: This data set was used for the identification of glucocorticoid regulated genes/transcripts in CCRF-CEM-C7H2 T-ALL cells.For each gene a representative transcript probe set was selected based on a score that combines average expression and variance expression across all samples. Differentially expressed genes/transcripts were identified for each time point by comparing expression values of GC treated to the respective control samples. Raw p-values were calculated employing the moderated t-test, and adjusted for multiple hypothesis testing with the method from Benjamini and Hochberg. Genes with an adjusted p-value smaller 0.05 and an absolute average M (log 2 fold change) value larger 0.7 were considered significantly differentially expressed.

Data type: Transcriptome or Gene expression

Sample scope: Multiisolate

Relevance: Medical

Release date: 2011-11-15

Last updated: 2011-05-02