Chromatin structure has long been suspected to have a role in regulating cell fate but there has been no convincing evidence for this. However, data published in Molecular Cell now support a model whereby transcription factors and epigenetic silencing are integrated quantitatively to control cell differentiation.
Hutchins and co-workers used helper-T-cell (TH cell) differentiation as a model system, and analysed TH cells from mice deficient in methyl-CpG-binding-domain protein 2 (Mbd2), one of the molecules that is thought to link DNA methylation and chromatin-based silencing.
In this study, cytokine signalling was manipulated to generate polarized TH1 and TH2 cells from uncommitted cells. The researchers found that, in Mbd2-deficient cells, the normal pattern of T-cell differentiation was dysregulated. For example, Mbd2-deficient cells produced ectopic interleukin (IL)-4 under TH1-polarizing conditions, and the normally essential transcriptional activator Gata-3 was dispensable for IL-4 induction. Moreover, expression of the signature genes encoding IL-4 and interferon γ was increased in TH2 and TH1 cells, respectively, suggesting that Mbd2 might be crucial in silencing these cytokine genes during TH-cell differentiation.
To investigate the relationships between Gata-3 and Mbd2, and the IL-4 gene, the authors determined relative Gata-3 messenger-RNA levels using the reverse-transcription polymerase chain reaction. Gata-3 mRNA levels were repressed as normal in cells cultured under TH1 conditions regardless of the Mbd2 genotype, indicating that it is not an excess of Gata-3 that induces IL-4 in Mbd2-mutant TH1 cells. In line with this observation, the authors proposed a quantitative competition model for activators (such as Gata-3) and silencers (such as Mbd2) whereby each regulator has an independent, quantitative effect.
To test this hypothesis, cells with three different Mbd2 gene doses (Mbd2+/+, Mbd2+/−, Mbd2−/−) were cultured under TH1 conditions (that is, where Gata-3 is not produced) and transduced with a bicistronic retrovirus encoding Gata-3 and green-fluorescent protein (GFP). Cells with different levels of Gata-3 were evaluated based on the level of green fluorescence intensity. Increasing levels of Gata-3 rendered cells competent for IL-4 production in a manner that is inversely correlated with Mbd2 gene dosage.
To investigate the basis for the competition between Gata-3 and Mbd2 further, the authors performed chromatin immunoprecipitation experiments. They found that Gata-3 seems to antagonize the binding of Mbd2 before demethylation of the IL-4 gene. This suggests that one of the key functions of a developmental transactivator might be to compete with silencing forces to reconfigure the activity of a target locus.
There are some indications that Gata-3 might have other, Mbd2-independent, roles in activating IL-4 expression. Future investigation might thus uncover further complexities in transcriptional programming in development.
References
ORIGINAL RESEARCH PAPER
Hutchins, A. S. et al. Gene silencing quantitatively controls the function of a developmental trans-activator. Mol. Cell 10, 81–91 (2002)
FURTHER READING
Bird, A. DNA methylation patterns and epigenetic memory. Genes Dev. 16, 6–21 (2002)
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Heinrichs, A. Competing forces. Nat Rev Mol Cell Biol 3, 632 (2002). https://doi.org/10.1038/nrm912
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DOI: https://doi.org/10.1038/nrm912