Translational profiling in childhood acute lymphoblastic leukemia: no evidence for glucocorticoid regulation of mRNA translation

Abstract

Background: Glucocorticoids (GCs) are natural stress induced steroid hormones causing cell cycle arrest and cell death in lymphoid tissues. Therefore they are the central component in the treatment of lymphoid malignancies, in particular childhood acute lymphoblastic leukemia (chALL). GCs act mainly via regulating gene transcription, which has been intensively studied by us and others. GC control of mRNA translation has also been reported but has never been assessed systematically. In this study we investigate the effect of GCs on mRNA translation on a genome-wide scale.

Results: Childhood T- (CCRF-CEM) and precursor B-ALL (NALM6) cells were exposed to GCs and subjected to "translational profiling", a technique combining sucrose-gradient fractionation followed by Affymetrix Exon microarray analysis of mRNA from different fractions, to assess the translational efficiency of the expressed genes. Analysis of GC regulation in ribosome-bound fractions versus transcriptional regulation revealed no significant differences, i.e., GC did not entail a significant shift between ribosomal bound and unbound mRNAs.

Conclusions: In the present study we analyzed for the first time possible effects of GC on the translational efficiency of expressed genes in two chALL model systems employing whole genome polysome profiling. Our results did not reveal significant differences in translational efficiency of expressed genes thereby arguing against a potential widespread regulatory effect of GCs on translation at least in the investigated in vitro systems.

Figure 1

Representative example of Agilent gel electrophoresis of RNA fractions obtained by sucrose gradient separation from NALM6 cells. Fractions 1–6 (pool 1) encompass non-ribosome bound RNAs, as suggested by the complete absence of 28S RNA. Fractions 7–8 form an intermediate pool potentially containing translationally-initiated mRNAs (pool 2). Fractions 9–20 (pool 3) contain mRNAs bound to ribosomes, as evidenced by the presence of both ribosomal subunits 18S and 28S.

Figure 2

Distribution of biotypes for the genes with the highest relative expressions in the corresponding pool. Distribution of biotypes in pools 1 (left panel) and 3 (middle panel) for the top 1% of genes with the highest relative expression in the corresponding pool compared to the total number of genes on the microarray (right panel). Shown are the results for EtOH-treated C7H2 cells. For the complete list of biotypes of the genes detectable on the microarray, see Table 1.

Figure 3

Distributions of translational efficiencies of protein-coding genes. Distributions of translational efficiencies of all protein-coding genes expressed as proportions of the gene mRNA in pool 3 for CEM-C7H2 (left panel) and NALM6 (right panel) cells. Red and blue color corresponds to GC- and EtOH-treated samples, respectively. Dotted lines indicate the median values.

Figure 4

Expression levels of mRNA for representative genes with high (A) and low (B) translational efficiencies. X-axis indicates 3 pools: non-translated (pool 1), intermediate (pool 2) and ribosome-bound (pool 3). Y-axis represents the expression level in log2 scale. Each dot corresponds to the average expression in 3 biological experiments, error bars show standard deviation. Values for GC- and EtOH-treated samples are drawn in red and blue, respectively. CEM-C7H2 cell line is indicated by solid lines, and NALM6 by dashed lines.