A switch and a dial

Study by the Schulz lab describes a novel regulatory principle involved in X-chromosome inactivation during development

In female mammalian cells, one of the two X chromosomes is deactivated to prevent the dangerous double expression of genes. This mechanism, driven by a gene called Xist, is responsible for several phenomena, ranging from the color patterning of certain cats' fur to color blindness and diseases in humans. Edda Schulz's lab has now together with the Vingron lab, identified novel regulators of Xist and discovered a two-step process that controls the network of gene regulatory elements that activate and regulate Xist during development.

In 1948, scientist William Barr and his graduate student observed unusual shapes in the nuclei of female cat nerve cells that were absent in male cats. These masses were later identified as inactive X chromosomes and named Barr bodies. They are the result of a process called X-inactivation. During the development of female mammals, one X chromosome is deactivated in each cell to compensate for the double expression of genes, which would otherwise be detrimental. This memory is retained in adult cells as the organism develops. Consequently, female mammals resemble more a mosaic of cells. In certain cats, for example, this leads to their characteristic color patterns, depending on which X chromosome is active and the color it codes for.

X-inactivation requires cells to sense the presence of two X chromosomes and randomly choose one for deactivation, steps that are conceptually often perceived as seperate. "It has become clear that these processes are not independent but rather part of a regulatory network that integrates both steps to inactivate one X chromosome," explains group leader Edda Schulz. "Our study could fill a major blind spot by identifying the regulators involved in this network and how they interact."

Many developmental genes, such as Xist, are regulated by elements close to the genes as well as by transcription factors often expressed from distant locations. “A few years ago, we mapped the nearby, or cis-regulatory, elements. Now, we wanted to figure out how these elements interact with transcription factors,” explains Till Schwämmle, the study's first author. To this end, the team developed an improved screening approach based on the CRISPR/Cas9 gene-editing tool. Using this tool, they identified novel regulatory factors and linked their interactions with other regulatory elements.

“We found that the Xist locus is regulated by two steps. In the first step a group of transcription factors that are preferetially expressed in females initiates what is essentially an on/off switch, making a decision which chromosome is deactivated. A second group of transcription factors is expressed later - and resembling a dial - boosts expression levels of Xist. This ensures proper silencing of the chromosome”, explains Till Schwämmle. 

One major remaining question is why Xist is only expressed in female cells with two X chromosomes. “We are now very interested in one of the factors Till discovered in his screen, called ZIC3 that drives this on-off switch”, says Edda Schulz. As it is encoded on the X-chromosomes, it has higher levels in female cells and could be a potential driver of female specific expression of Xist. “We and others have been long looking for such X-linked activators and will now further investigate their roles”, she concludes.