In one of the lower valleys of Waddington’s epigenetic landscape, T cells are confronted with a fatal decision: Become a CD4+ helper T cell and assist your brethren in fighting evil invaders or turn into a CD8+ cytotoxic T cell and do the dirty work. But as in all good fantasy novels, our heroes first have to undergo a series of trials to prove their worthiness.
Early CD4−CD8− double-negative (DN) progenitor cells transition through a series of stages that ultimately leads to the random rearrangement of their T cell antigen receptors (TCRs) and the upregulation of CD4 and CD8 expression. These CD4+CD8+ double-positive (DP) progenitors then test their TCRs for specificity for major histocompatibility complex (MHC) class II or MHC class I molecules, ultimately determining whether they may enter the helper (CD4+) or the cytotoxic (CD8+) lineage.
Intrigued by this fantastic process, a crafty fellowship of researchers led by Dan R Littman set out to study the epigenetic processes controlling CD4 expression during T cell lineage choice. Prior studies had already identified a silencer (S4) and a proximal enhancer (E4P) as critical cis-acting elements that control the initiation but not the maintenance of Cd4 repression and expression, respectively. Based on these findings the researchers designed a retroviral shRNA screen for the identification of trans-acting factors that are involved in Cd4 repression in CD8+ cytotoxic T cells.
Here is what they found:
- 83% of shRNAs that caused an upregulation of CD4 in CD8+ T cells were specific for the DNA methyltransferase Dnmt1
- This result was confirmed in T cells carrying a hypomorphic mutation in the Dnmt1 locus
- DNA methylation analysis revealed a differentially methylated region (DMR) that is hypermethylated in CD8+ and hypomethylated in CD4+ cells
- Surprisingly, hypermethylation of the DMR was already present at the DP stage
- The S4 silencer mediated activity was required for sustaining Cd4 repression during transition of DP to CD8+ cells but was not needed to maintain locus methylation in DP cells
- Deletion of the E4P enhancer sequence resulted in hypermethylation and unstable Cd4 expression in T helper cells
- During commitment to the T helper lineage the DMR was demethylated in E4P-dependent manner
- The DNA demethylation of the Cd4 DMR was cell division-independent and correlated with increased 5-hmC levels suggesting an active TET enzyme-dependent pathway
With this, the researchers conclude that DNA demethylation is critical for heritable expression and that the Cd4 locus represents an ideal model to study the mechanisms controlling this crucial process.
To read all the helpful details head over to Nature Immunology, June 2015.