TY - JOUR

T1 - Efficient Pre-mRNA Cleavage Prevents Replication-Stress-Associated Genome Instability

AU - Teloni, Federico

AU - Kilic, Sinan

AU - Menon, Shruti

AU - Imhof, Ralph

AU - Janscak, Pavel

N1 - Funding Information: We are grateful to ScopeM, ZMB, and the FGCZ for excellent support. We thank R. Crouch for the RNaseH1 construct, E. Wahle for WDR33 cDNA, E. Soutoglou for GFP-LacI and GFP-LacI-ΔEMD cells, R. Greenberg for U-2 OS 2-6-3 cells, and M. Lopes and M. Berti for sharing protocols and reagents. We thank all members of our labs and of the DMMD for discussions and advice. Research in the lab of M.A. is supported by the Swiss National Science Foundation (SNSF, 150690 and 179057), the European Research Council (ERC) under the European Union's Horizon 2020 research and innovation program (ERC-2016-STG 714326), the Novartis Foundation for Medical-Biological Research (Grant 16B078), and the Swiss Foundation to Combat Cancer (Stiftung zur Krebsbekämpfung). J.M. is supported by the Gobierno Vasco Programa Posdoctoral de Perfeccionamiento de Personal Investigador Doctor. F.T. received support from the UZH Candoc Program (FK-16-053). Research in the lab of P.J. is supported by grants from the SNSF (31003A_166451), Swiss Cancer League (KFS-3802-02-2016), and the Czech Science Foundation (17-02080S), as well as the Neuron Fund for Support of Science and European Regional Development Fund/OP RDE (CZ.02.1.01/0.0/0.0/16_013/0001775) to J.D. Research in the lab of T.B. is supported by the SNSF (157488 and 180345). Funding Information: We are grateful to ScopeM, ZMB, and the FGCZ for excellent support. We thank R. Crouch for the RNaseH1 construct, E. Wahle for WDR33 cDNA, E. Soutoglou for GFP-LacI and GFP-LacI-ΔEMD cells, R. Greenberg for U-2 OS 2-6-3 cells, and M. Lopes and M. Berti for sharing protocols and reagents. We thank all members of our labs and of the DMMD for discussions and advice. Research in the lab of M.A. is supported by the Swiss National Science Foundation (SNSF, 150690 and 179057 ), the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation program (ERC-2016-STG 714326 ), the Novartis Foundation for Medical-Biological Research (Grant 16B078 ), and the Swiss Foundation to Combat Cancer (Stiftung zur Krebsbekämpfung). J.M. is supported by the Gobierno Vasco Programa Posdoctoral de Perfeccionamiento de Personal Investigador Doctor . F.T. received support from the UZH Candoc Program ( FK-16-053 ). Research in the lab of P.J. is supported by grants from the SNSF ( 31003A_166451 ), Swiss Cancer League ( KFS-3802-02-2016 ), and the Czech Science Foundation ( 17-02080S ), as well as the Neuron Fund for Support of Science and European Regional Development Fund/OP RDE ( CZ.02.1.01/0.0/0.0/16_013/0001775 ) to J.D. Research in the lab of T.B. is supported by the SNSF ( 157488 and 180345 ). Publisher Copyright: © 2018 The Author(s)

PY - 2019/2/21

Y1 - 2019/2/21

N2 - Cellular mechanisms that safeguard genome integrity are often subverted in cancer. To identify cancer-related genome caretakers, we employed a convergent multi-screening strategy coupled to quantitative image-based cytometry and ranked candidate genes according to multivariate readouts reflecting viability, proliferative capacity, replisome integrity, and DNA damage signaling. This unveiled regulators of replication stress resilience, including components of the pre-mRNA cleavage and polyadenylation complex. We show that deregulation of pre-mRNA cleavage impairs replication fork speed and leads to excessive origin activity, rendering cells highly dependent on ATR function. While excessive formation of RNA:DNA hybrids under these conditions was tightly associated with replication-stress-induced DNA damage, inhibition of transcription rescued fork speed, origin activation, and alleviated replication catastrophe. Uncoupling of pre-mRNA cleavage from co-transcriptional processing and export also protected cells from replication-stress-associated DNA damage, suggesting that pre-mRNA cleavage provides a mechanism to efficiently release nascent transcripts and thereby prevent gene gating-associated genomic instability. Replication stress is a hallmark of many cancers. Teloni et al. identify the pre-mRNA cleavage factor WDR33 as regulator of replication stress resilience and demonstrate that, when WDR33 function is impaired, unreleased nascent transcripts and genomic loci re-localize toward the nuclear periphery, where they cause replication stress and DNA damage.

AB - Cellular mechanisms that safeguard genome integrity are often subverted in cancer. To identify cancer-related genome caretakers, we employed a convergent multi-screening strategy coupled to quantitative image-based cytometry and ranked candidate genes according to multivariate readouts reflecting viability, proliferative capacity, replisome integrity, and DNA damage signaling. This unveiled regulators of replication stress resilience, including components of the pre-mRNA cleavage and polyadenylation complex. We show that deregulation of pre-mRNA cleavage impairs replication fork speed and leads to excessive origin activity, rendering cells highly dependent on ATR function. While excessive formation of RNA:DNA hybrids under these conditions was tightly associated with replication-stress-induced DNA damage, inhibition of transcription rescued fork speed, origin activation, and alleviated replication catastrophe. Uncoupling of pre-mRNA cleavage from co-transcriptional processing and export also protected cells from replication-stress-associated DNA damage, suggesting that pre-mRNA cleavage provides a mechanism to efficiently release nascent transcripts and thereby prevent gene gating-associated genomic instability. Replication stress is a hallmark of many cancers. Teloni et al. identify the pre-mRNA cleavage factor WDR33 as regulator of replication stress resilience and demonstrate that, when WDR33 function is impaired, unreleased nascent transcripts and genomic loci re-localize toward the nuclear periphery, where they cause replication stress and DNA damage.

KW - ATR

KW - checkpoint activation

KW - cleavage

KW - gene gating

KW - origin firing

KW - polyadenylation

KW - pre-mRNA processing

KW - R-loops

KW - replication catastrophe

KW - replication stress

KW - RNA:DNA hybrids

UR - http://www.scopus.com/inward/record.url?scp=85061541528&partnerID=8YFLogxK

U2 - 10.1016/j.molcel.2018.11.036

DO - 10.1016/j.molcel.2018.11.036

M3 - Journal article

C2 - 30639241

AN - SCOPUS:85061541528

VL - 73

SP - 670-683.e12

JO - Molecular Cell

JF - Molecular Cell

SN - 1097-2765

IS - 4

ER -