• A non-coding RNA regulates genomic G4 structures–Nov 2022
• Owen Shen won the first place award in the student oral presentation held by The Taiwan Society for Biochemistry and Molecular Biology–Nov 2022.
• XPF induces breaks for Alternative Lengthening of Telomeres—Oct 2022

TERRA regulates DNA G-quadruplex formation and ATRX recruitment to chromatin

Published in Nucleic Acids Research, Nov 2022

https://academic.oup.com/nar/advance-article/doi/10.1093/nar/gkac1114/6849496

The genome consists of non-B-DNA structures such as G-quadruplexes (G4) which are enriched at active chromatin and nucleosome-depleted regions. TERRA is capable of folding into an RNA G-quadruplex and interacting with chromatin remodeler ATRX. In this article, we uncover that TERRA regulates the DNA G-quadruplex (G4) structures and ATRX occupancy near transcription start sites in vivo.  

TERRA depletion leads to increased ATRX occupancy on repetitive sequences and subtelomeric regions. Strikingly, ATRX ectopically binds to TSS in TERRA knockdown cells. Such elevation of ATRX loading at transcription start sites is accompanied by the reduction of DNA G4 structures and gene repression. Loss of ATRX alleviates the effect of gene repression caused by TERRA depletion. Our findings provide insights into the mechanism by which TERRA regulates gene expression in cis and in trans.

We propose that the properties of G-rich sequences in TERRA RNA can sequester G4 binding proteins and preserve DNA G4 structures. This is the first study showing that a long non-coding RNA is capable of regulating DNA G4 across the genome.

XPF activates break-induced Telomere synthesis

https://www.nature.com/articles/s41467-022-33428-0

The story depicted that TERRA R-loops and XPF are pivotal in the alternative lengthening of telomeres (ALT) pathway. TERRA is a long non-coding RNA, which contains telomeric repeat sequences and forms DNA:RNA hybrids (called TERRA R-loops) at telomeres. TERRA R-loops are specifically enriched at telomeres in cancer cells utilizing the ALT mechanism, which relies on homologous recombination and break-induced telomere synthesis to extend telomere length independent of telomerase. Although it is known that DNA repair is involved in the ALT mechanism, it is unclear how cells initiate ALT and what contexts that trigger DNA damage response at telomeres in ALT cancer. In this study, we uncovered that TERRA R-loops and XPF are required for the onset of ALT. TERRA R-loops trigger telomere clustering and activate DNA damage response by recruiting XPF. Such DNA damage response at telomeres is required for inducing homologous recombination and telomere synthesis in ALT cancer cells.

We developed an RCas9 system to specifically deplete TERRA RNA without editing telomeric DNA in ALT cells and found that TERRA depletion reduces alternative lengthening of telomeres. TERRA iDRiP-MS in ALT cells revealed that TERRA interacts with a large subset of proteins involved in DNA repair pathway. Interestingly, TERRA particularly interacts with several nucleotide excision repair (NER) factors including XPF in ALT cells. Our results show that TERRA R-loops recruit the endonuclease XPF to telomeres, leading to DNA double-strand breaks to activate break-induced telomere synthesis at ALT telomeres.

Targeting XPF by small interference inhibits cell growth in ALT cancer cells and reduces telomere lengthening. These findings may provide insights into ALT cancer therapy.