Exploring the mechanisms governing nuclear cGAS localization and activity

Recognition of pathogen-derived molecules through germline-encoded receptors is a fundamental principle of innate immunity. Pattern recognition receptors detect specific intracellular danger signals to trigger potent immune responses. The DNA sensor cyclic GMP-AMP synthase (cGAS) detects double-stranded DNA derived from viruses and microbes in the cytoplasm. However, due to its ability to recognize DNA independent of sequence, aberrant detection of self-DNA in various settings can lead to autoinflammatory diseases. Under physiological conditions, genomic DNA is sequestered in the nucleus and shielded from cytosolic molecules by the nuclear envelope (NE). Nevertheless, cGAS gains access to the nuclear compartment in some instances, and the mechanisms controlling its activity on genomic DNA are not fully known.In this study, we investigate the regulation of nuclear cGAS localization and activity in different contexts. We first describe how cGAS activity against nuclear DNA is restricted in instances of spontaneous loss of nucleo-cytoplasmic compartmentalization. Upon NE rupture, cGAS enters the nucleus, but is prevented from activation by Barrier-to-autointegration factor 1 (BAF). BAF dynamically competes with cGAS for DNA binding, thereby preventing the formation of stable cGAS-DNA complexes and cGAS enzymatic activity. We thus identify a cellular safeguard mechanism, beyond physical separation, that restricts cGAS activity against genomic self-DNA.In addition, we investigate cGAS activation in conditions associated with a disrupted NE. Instability of the NE is a hallmark of laminopathies, diseases of the nuclear lamina that can manifest in premature aging phenotypes. While we find no contribution of basal cGAS signaling to the pathogenesis of two different progeria syndromes, we detect a decreased capacity to respond to DNA challenge on the cellular level, suggesting that cGAS may be sequestered inside the nucleus in these cells.Lastly, because cGAS is increasingly recognized to reside inside the nucleus at steady state, we delineate cGAS nuclear localization in time and space. In cycling cells, nuclear cGAS levels oscillate with the cell cycle and are highest after mitosis followed by a sharp decrease in G1 phase. We report that cGAS is not exported during G1, but rather subjected to degradation inside the nucleus by the nuclear proteasome. Moreover, we establish a technique to study potential interaction partners of nuclear cGAS, which opens up intriguing possibilities for further investigation.Together, our findings contribute to a better understanding of the regulatory processes that govern nuclear cGAS localization and activity, and will help to uncover potential disease-causing aberrations in the future.