DNA G-quadruplex structures in the mammalian genome : dissecting genome-wide mechanisms of formation and turnover of DNA G-quadruplexes

Since the beginning of 20th century, when the G-tetrad was first discovered, scientists have been seeking to understand the role of G4 quadruplex structures (G4s) in replication, DNA damage, telomere integrity and the regulation of gene expression. However, it is likely that the field has just scratched the tip of the iceberg when it comes to assessing and interpreting the function of individual G4s, forming in a specific locus, in the complex context of chromatin. This is partially due to the limitations of available genome-wide G4 mapping methods, for example in terms of sensitivity and signal-to-noise ratio. Therefore, to further investigate G4 dynamics in mammalian cells, it is critical to develop robust, sensitive and quantitative methods to map endogenous G4s genome-wide.

To achieve this, in paper I, we developed a quantitative method for genome-wide G4 mapping, termed G4 qCUT&Tag, by combining a G4-specific antibody with a modified CUT&Tag. We showed that G4 qCUT&Tag accurately and reproducibly generates genome-wide G4 maps, allowing us to study: 1) G4 landscapes in mESCs; 2) relation of G4s with other chromatin features in mESCs; 3) G4 dynamics during transcription and mESCs differentiation; (4) correlation of G4s and R-loops.

G4s largely correlate with open chromatin, and since nucleosomes and G4s are mutually exclusive on a given stretch of DNA, it is believed that nucleosome dynamics affect G4 dynamics and vice versa. Thus, in paper II, we elucidated the mechanism of histone turnover and histone variant H3.3 incorporation in mESC-specific interstitial heterochromatin. We discovered that in the H3K9me3-featured heterochromatin the chromatin remodeler Smarcad1 evicts nucleosome and create accessible DNA, allowing H3.3 incorporation. The proposed model in paper II provided the basis for understanding the causal relationship between G4s and H3.3 incorporation in the heterochromatin.

List of scientific papers

I. Jing Lyu, Rui Shao, Philip Yuk Kwong Yung, Simon J. Elsässer. Genome-wide mapping of G-quadruplex structures with CUT&Tag. Nucleic Acids Research. 2021 Nov 18.
https://doi.org/10.1093/nar/gkab1073

II. Carmen Navarro, Jing Lyu*, Anna-Maria Katsori*, Rozina Caridha, Simon J. Elsässer. An embryonic stem cell-specific heterochromatin state promotes core histone exchange in the absence of DNA accessibility. Nature Communications. 2020 Oct 9;11(1):5095. *These authors contributed equally.
https://doi.org/10.1038/s41467-020-18863-1