ANNULÉ - Philippe Pasero - Cellular responses to replication stress: from R-loops to inflammation and beyond - IPBS-Toulouse, Seminar Room

Dr. Philippe Pasero

IGH, Montpellier, France

Cellular responses to replication stress: from R-loops to inflammation and beyond

Replication stress (RS) commonly refers to alterations of fork progression caused by various events of endogenous or exogenous origin. Oncogenes such as RASV12 induce RS by increasing transcription-replication conflicts (TRCs), but the mechanisms involved remain poorly understood. TRCs could result from the presence of structures called R-loops that form during transcription and may act as roadblocks to replication. Alternatively, toxic R-loops could form as a consequence of fork arrest and interfere with replication restart. To discriminate between these two possibilities, we have analyzed fork arrest and restart in the absence of RNase H activity. These analyses showed that yeast and human cells fail to process and to restart arrested forks in the presence of persistent RNA:DNA hybrids. We propose that cotranscriptional R-loops are converted into toxic RNA:DNA hybrids upon fork passage, which interfere with fork restart. The expression of RASV12 in immortalized BJ fibroblasts promotes oncogene-induced senescence (OIS) by increasing RS and triggering inflammation. Interestingly, BJ cells adapt to RASV12-induced RS by overexpressing fork-associated components of the ATR-CHK1 pathway, such as Claspin and Timeless. Interestingly, these cells show a reduction in both RS and inflammation, suggesting that these processes are linked. To address this possibility, we have inhibited MRE11 in BJ-RASV12 cells and found that it completely suppressed OIS. Since MRE11 releases DNA fragments from stressed forks, these data suggest that MRE11 connects RS, inflammation and senescence by promoting the accumulation of immunogenic DNA fragments. This view is supported by the fact that inhibition of TREX1, the main exonuclease involved in the degradation of cytosolic DNA, is sufficient to induce replication stress and senescence, even in the absence of oncogenic stress. Altogether, these data indicate that MRE11 plays a central role in OIS by linking the replication stress response to inflammation.

Selected references

• Promonet A, Padioleau I, Liu Y, Sanz L, Biernacka A, Schmitz AL, Skrzypczak M, Sarrazin A, Mettling C, Rowicka M, Ginalski K, Chedin F, Chen CL, Lin YL and Pasero P (2020) Topoisomerase 1 prevents replication stress at R-loop-enriched transcription termination sites. Nat Commun 11, 3940
• Forey R, Poveda A, Sharma S, Padioleau I, Renard C, Lambert R, Skrzypczak M, Ginalski K, Lengronne A, Chabes A, Pardo B and Pasero P (2020) Mec1 is activated at the onset of normal S phase by low dNTP pools impeding DNA replication. Mol Cell 78, 396-410
• Delamarre A, Barthe A, de la Roche Saint‐André C, Luciano P, Forey R, Padioleau I, Skrzypczak M, Ginalski K, Géli V, Pasero P and Lengronne A (2020) MRX increases chromatin accessibility at stalled replication forks to promote nascent DNA resection and cohesin loading. Mol Cell 77, 395-410
• Bianco JN, Bergoglio V, Lin YL, Pillaire MJ, Schmitz AL, Gilhodes J, Lusque A, Mazières J, Lacroix-Triki M, Roumeliotis TI, Choudhary J, Moreaux J, Hoffmann JS, Tourrière H and Pasero P (2019) Overexpression of Claspin and Timeless protects cancer cells from replication stress in a checkpoint-independent manner. Nat Commun 10, 910
• Bacal J, Moriel-Carretero M, Pardo B, Barthe A, Sharma S, Chabes A, Lengronne A and Pasero P (2018) Mrc1 and Rad9 cooperate to regulate initiation and elongation of DNA replication in response to DNA damage. EMBO J 37, e99319
• Coquel F, Silva MJ, Técher H, Zadorozhny K, Sushma S, Nieminuszczy J, Mettling C, Dardillac E, Barthe A, Schmitz AL, Promonet A, Cribier A, Sarrazin A, Niedzwiedz W, Lopez B, Costanzo V, Krejci L, Chabes A, Benkirane M, Lin YL and Pasero P (2018) SAMHD1 acts at stalled replication forks to prevent induction of interferon. Nature 557, 57-61