Transcription factors control gene expression programs in the context of the chromatin structure of their target genes. DNA methylation, post-translational histone modifications such as acetylation and methylation, and higher order chromatin organization allow the maintenance of gene expression patterns through mitosis, but how do they accommodate developmentally regulated changes in gene expression programs? Although histone acetylation and deacetylation are in dynamic equilibrium and mechanisms for the removal of methyl groups from histones are emerging, the extent to which there is active demethylation of DNA remains controversial. Looking at chromatin in the three-dimensional space of the nucleus, recent work demonstrates that gene regulation involves contacts between regulatory elements within genes or gene clusters on the same chromosome (in cis) and between different chromosomes (in trans). Finally, non-coding RNAs make a significant contribution to transcriptional and post-transcriptional gene silencing. Together, these advances contribute to an understanding of how gene expression programs are established, maintained and modified during development.