title: RNA-processing proteins regulate Mec1/ATR activation by promoting generation of RPA-coated ssDNA.
abstract: Eukaryotic cells respond to DNA double-strand breaks (DSBs) by activating a checkpoint that depends on the protein kinases Tel1/ATM and Mec1/ATR. Mec1/ATR is activated by RPA-coated single-stranded DNA (ssDNA), which arises upon nucleolytic degradation (resection) of the DSB. Emerging evidences indicate that RNA-processing factors play critical, yet poorly understood, roles in genomic stability. Here, we provide evidence that the Saccharomyces cerevisiae RNA decay factors Xrn1, Rrp6 and Trf4 regulate Mec1/ATR activation by promoting generation of RPA-coated ssDNA. The lack of Xrn1 inhibits ssDNA generation at the DSB by preventing the loading of the MRX complex. By contrast, DSB resection is not affected in the absence of Rrp6 or Trf4, but their lack impairs the recruitment of RPA, and therefore of Mec1, to the DSB. Rrp6 and Trf4 inactivation affects neither Rad51/Rad52 association nor DSB repair by homologous recombination (HR), suggesting that full Mec1 activation requires higher amount of RPA-coated ssDNA than HR-mediated repair. Noteworthy, deep transcriptome analyses do not identify common misregulated gene expression that could explain the observed phenotypes. Our results provide a novel link between RNA processing and genome stability.
authors.affiliation: Dipartimento di Biotecnologie e Bioscienze, Università di Milano-Bicocca, Milan, Italy.
authors.forename: Nicola
authors.initials: N
authors.lastname: Manfrini
authors.affiliation: Dipartimento di Biotecnologie e Bioscienze, Università di Milano-Bicocca, Milan, Italy.
authors.forename: Camilla
authors.initials: C
authors.lastname: Trovesi
authors.affiliation: Institut Curie, CNRS UMR3244 Université Pierre et Marie Curie, Paris Cedex 05, France.
authors.forename: Maxime
authors.initials: M
authors.lastname: Wery
authors.affiliation: Dipartimento di Biotecnologie e Bioscienze, Università di Milano-Bicocca, Milan, Italy.
authors.forename: Marina
authors.initials: M
authors.lastname: Martina
authors.affiliation: Dipartimento di Biotecnologie e Bioscienze, Università di Milano-Bicocca, Milan, Italy.
authors.forename: Daniele
authors.initials: D
authors.lastname: Cesena
authors.affiliation: Institut Curie, CNRS UMR3244 Université Pierre et Marie Curie, Paris Cedex 05, France.
authors.forename: Marc
authors.initials: M
authors.lastname: Descrimes
authors.affiliation: Institut Curie, CNRS UMR3244 Université Pierre et Marie Curie, Paris Cedex 05, France antonin.morillon@curie.fr fabrizio.dadda@ifom.eu mariapia.longhese@unimib.it.
authors.forename: Antonin
authors.initials: A
authors.lastname: Morillon
authors.affiliation: IFOM Foundation-FIRC Institute of Molecular Oncology Foundation, Milan, Italy Istituto di Genetica Molecolare, Consiglio Nazionale delle Ricerche, Pavia, Italy antonin.morillon@curie.fr fabrizio.dadda@ifom.eu mariapia.longhese@unimib.it.
authors.forename: Fabrizio
authors.initials: F
authors.lastname: d'Adda di Fagagna
authors.affiliation: Dipartimento di Biotecnologie e Bioscienze, Università di Milano-Bicocca, Milan, Italy antonin.morillon@curie.fr fabrizio.dadda@ifom.eu mariapia.longhese@unimib.it.
authors.forename: Maria Pia
authors.initials: MP
authors.lastname: Longhese
mesh_terms: D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair / Q000235:genetics / Q000502:physiology; D004277:DNA, Single-Stranded / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004259:DNA-Directed DNA Polymerase / Q000235:genetics / Q000378:metabolism; D005095:Exoribonucleases / Q000235:genetics / Q000378:metabolism; D063326:Exosome Multienzyme Ribonuclease Complex / Q000235:genetics / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D051716:Replication Protein A / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; D051716:Replication Protein A; D029701:Saccharomyces cerevisiae Proteins; C088937:MEC1 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases; D004259:DNA-Directed DNA Polymerase; C487545:PAP2 protein, S cerevisiae; D005095:Exoribonucleases; D063326:Exosome Multienzyme Ribonuclease Complex; C068398:XRN1 protein, S cerevisiae; C112610:RRP6 protein, S cerevisiae
keywords: DNA damage checkpoint; DNA double‐strand breaks; Rrp6; Trf4; Xrn1