High-throughput engineering of a mammalian genome reveals building principles of methylation states at CG rich regions

Regions of DNA called genes produce the proteins and other molecules that are essential for life. The act of making these molecules is known as gene expression, and being able to switch this process on and off allows cells to adapt to changing conditions. For example, some genes may be turned on in response to injury or may only turn on during waking hours. There are several ways gene expression can be switched on and off. Proteins called transcription factors can bind to DNA and act like a switch that affects nearby genes. Alternatively, special tags called methyl groups can attach to the ‘letters’ that make up the DNA code and turn off gene expression. However, it is not understood how these tags work with transcription factors and other forms of gene regulation. Regions of DNA that boost the expression of a neighboring gene are called promoters. Many promoters in mammals contain repeating patterns of the DNA letters ‘C’ (which is a chemical called cytosine) and ‘G’ (guanine), and these regions are tagged less often than other regions of DNA. This led scientists to wonder whether the DNA sequence itself controls where the tags are placed, but existing experimental techniques made it difficult to establish if DNA sequence alone can prevent tagging. Krebs et al. created a technique that allows thousands of different DNA sequences to be inserted into the same part of the genome of mouse stem cells. Comparing the tagging across these different sequences revealed that the CG pattern is not as closely associated with tagging as was thought. If the CG pattern is repeated many times it does seem to prevent tagging, but sequences with fewer repeats also sometimes escape tagging. Krebs et al. found that a sequence was much less likely to be tagged if the nearby DNA also contains a site that transcription factors can bind to. However, regions with a very high number of CG repeats are able to avoid tagging without help from transcription factors. Krebs et al. found that this behavior is not seen in cancer cells. DNA in cancer cells is heavily tagged, even in CG-rich regions, and transcription factors do not appear to play a major role in directing tagging. The new approach developed by Krebs et al. should benefit researchers working to understand the multiple mechanisms that control gene activity.