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H2020Individual fellowship2021–2024

GLoopID · Mechanisms underlying regulation and removal of G-quadruplex/R-loop transcription-replication conflicts

Horizon 2020 — Marie Skłodowska-Curie Actions

Duration
2021-09-01 → 2024-01-04
EU contribution
€224,934
Participants
1
Scheme
MSCA-IF

Lines connect the coordinator with its partners.

Results in brief

Mechanisms underlying regulation and removal of G-quadruplex/R-loop transcription-replication conflicts

Guanine-rich repetitive DNA sequences, which can form four-stranded structures (G-quadruplex or G4), preferentially form in single stranded DNA (ssDNA) upon transient melting and can also occur in RNA. Transcribed G4s often co-exist with stable DNA:RNA hybrids forming a G4/R-loop, potentially causing deleterious transcription-replication conflicts. G4 motifs are enriched at regulatory loci such as active promoters or enhancers and at telomeres, and they have been implicated in several key biological processes, such as DNA replication, transcription, and telomere homeostasis. Telomeric G4/R-loops are linked to cancers that maintain telomere length not by upregulating telomerase but instead using a recombination-based telomere maintenance mechanism called alternative lengthening of telomeres (ALT). ALT constitutes a pathological scenario where aberrant telomeric chromatin triggers G4/R-loops that drives telomeric replication stress and recombination. ALT cancers are a major cancer type of unmet clinical need: typically mesenchymal in origin, with poor clinical outcomes and account for approximately 10-15% of all tumours. Whereas ALT tumours are strongly associated with mutations in the chromatin remodeller ATRX, inactivation of ATRX alone in culture is insufficient for a cell to become ALT. • Specific Aim 1: Determine the proximity proteome composition of site-specific G4/R-loop TRCs (WP1) • Specific Aim 2: Characterise the genetic vulnerabilities of G4/R-loop helicase-deficient cells (WP2) • Specific Aim 3: Identify the protein domains of G4/R-loop helicases essential for TRC removal (WP3)

Data: CORDIS, © European Union

Project objective

Maintenance of genome stability is challenged by obstacles that interfere with normal progression of essential DNA-associated transactions, such as DNA replication and transcription. One such obstacle is the G-quadruplex (G4) DNA secondary structure, which can form in G-rich repetitive DNA sequences. Transcribed G4-DNA loci often co-exist with stable RNA-DNA hybrids (R-loops), potentially causing deleterious transcription-replication conflicts (TRCs). Therefore, the ability of cells to mitigate TRCs formation is critical for cell and organismal fitness. While the pathways involved in repairing other DNA lesions are relatively well understood, the mechanisms by which cells respond to G4/R-loop-induced TRCs remain uncharted, largely due to a lack of efficient approaches for inducing site-specific G4s/R-loops into the genome. Several studies reported dedicated helicases capable of unwinding G4s/R-loops, yet whether additional factors exist and how different types of TRCs are resolved remains unknown. In this project, I aim to dissect the fundamental mechanisms that protect cells from TRCs at DNA loci harbouring G4s/R-loops. To do so, I will establish a novel system with site-specific G4/R-loop substrates in living cells and I will systematically investigate their protein composition to identify novel factors in G4/R-loop metabolism. I will further examine the genetic vulnerabilities of candidate G4/R-loop helicase-deficient cells and the regulatory mechanisms underlying G4/R-loop unwinding by these helicases. As mutations in several G4/R-loop helicases predispose to various cancers and genetic disorders, a detailed understanding of the basic mechanisms controlling G4/R-loop removal is of utmost importance, and will shed light on disease aetiology and the rational development of more targeted therapeutic strategies.

Original text from CORDIS.

Participants

  • THE FRANCIS CRICK INSTITUTE LIMITED · LondonCoordinatorUnited Kingdom

Links

Data: CORDIS, © European Union