Cyclic GMP–AMP synthase (cGAS) is a sensor for double-stranded DNA, activation of which leads to synthesis of the second messenger cyclic GMP–AMP (cGAMP) and ultimately to the production of type I interferons and pro-inflammatory cytokines. In the case of microbial infection, this is an important protective component of innate immunity, but in the case of exposure to self-DNA, it can result in auto-inflammation and pathology. A particular point of self-susceptibility occurs during mitosis when the nuclear envelope breaks down and cGAS is exposed to nuclear DNA. Chromatin association through nucleosome tethering has previously been shown to partially inhibit cGAS activity; here, Zhijian Chen and colleagues describe an additional layer of cGAS regulation during the danger zone of mitosis.
Further experiments showed that cGAS is phosphorylated at six serine residues and two threonine residues within the amino terminus when HeLa cells and primary human dermal fibroblasts enter M phase. The authors proposed that these negatively charged phosphorylated residues might interfere with the ionic interactions that otherwise occur between positively charged lysine and arginine residues in the N terminus of cGAS and DNA. Indeed, when all of the N-terminal serine and threonine residues were replaced with aspartate and glutamate, respectively, to mimic the electrostatic effect of phosphorylation, the activity of the mutant cGAS (cGAS20DE) was largely abolished. By contrast, HeLa cGAS-knockout cells expressing cGAS20A (in which all serine and threonine residues are replaced with alanine to prevent phosphorylation) produced significantly more cGAMP than knockout cells expressing wild-type cGAS. cGAS with mutation of the nucleosome-tethering sites Arg236 or Arg255 was active in asynchronous cell lysates containing chromosomal DNA but inhibited in mitotic cell lysates; additional cGAS20A mutation restored cGAS activity in mitotic lysates. Thus, phosphorylation and nucleosome tethering function independently to inhibit cGAS during mitosis.
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