title: Altered spatio-temporal dynamics of RNase H2 complex assembly at replication and repair sites in Aicardi-Goutières syndrome.
abstract: Ribonuclease H2 plays an essential role for genome stability as it removes ribonucleotides misincorporated into genomic DNA by replicative polymerases and resolves RNA/DNA hybrids. Biallelic mutations in the genes encoding the three RNase H2 subunits cause Aicardi-Goutières syndrome (AGS), an early-onset inflammatory encephalopathy that phenotypically overlaps with the autoimmune disorder systemic lupus erythematosus. Here we studied the intracellular dynamics of RNase H2 in living cells during DNA replication and in response to DNA damage using confocal time-lapse imaging and fluorescence cross-correlation spectroscopy. We demonstrate that the RNase H2 complex is assembled in the cytosol and imported into the nucleus in an RNase H2B-dependent manner. RNase H2 is not only recruited to DNA replication foci, but also to sites of PCNA-dependent DNA repair. By fluorescence recovery after photobleaching, we demonstrate a high mobility and fast exchange of RNase H2 at sites of DNA repair and replication. We provide evidence that recruitment of RNase H2 is not only PCNA-dependent, mediated by an interaction of the B subunit with PCNA, but also PCNA-independent mediated via the catalytic domain of the A subunit. We found that AGS-associated mutations alter complex formation, recruitment efficiency and exchange kinetics at sites of DNA replication and repair suggesting that impaired ribonucleotide removal contributes to AGS pathogenesis.
authors.affiliation: Department of Pediatrics, Medizinische Fakultät Carl Gustav Carus and.
authors.forename: Barbara
authors.initials: B
authors.lastname: Kind
authors.affiliation: Department of Biology, Technische Universität Darmstadt, 64287 Darmstadt, Germany.
authors.forename: Britta
authors.initials: B
authors.lastname: Muster
authors.affiliation: Biotechnology Center, Technische Universität Dresden, 01307 Dresden, Germany.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Staroske
authors.affiliation: Department of Physics, Applied Physics and Astronomy, Rensselaer Polytechnic Institute, New York 12180-3590, USA and.
authors.forename: Henry D
authors.initials: HD
authors.lastname: Herce
authors.affiliation: Institute of Earth and Environmental Science, Potsdam University, 14476 Potsdam, Germany.
authors.forename: René
authors.initials: R
authors.lastname: Sachse
authors.affiliation: Department of Biology, Technische Universität Darmstadt, 64287 Darmstadt, Germany.
authors.forename: Alexander
authors.initials: A
authors.lastname: Rapp
authors.affiliation: Department of Pediatrics, Medizinische Fakultät Carl Gustav Carus and.
authors.forename: Franziska
authors.initials: F
authors.lastname: Schmidt
authors.affiliation: Department of Pediatrics, Medizinische Fakultät Carl Gustav Carus and.
authors.forename: Sarah
authors.initials: S
authors.lastname: Koss
authors.affiliation: Department of Biology, Technische Universität Darmstadt, 64287 Darmstadt, Germany, minae.lee-kirsch@uniklinikum-dresden.de cardoso@bio.tu-darmstadt.de.
authors.forename: M Cristina
authors.initials: MC
authors.lastname: Cardoso
authors.affiliation: Department of Pediatrics, Medizinische Fakultät Carl Gustav Carus and minae.lee-kirsch@uniklinikum-dresden.de cardoso@bio.tu-darmstadt.de.
authors.forename: Min Ae
authors.initials: MA
authors.lastname: Lee-Kirsch
mesh_terms: D020274:Autoimmune Diseases of the Nervous System / Q000201:enzymology* / Q000235:genetics; D002467:Cell Nucleus / Q000201:enzymology / Q000235:genetics; D003600:Cytosol / Q000201:enzymology; D004249:DNA Damage; D004261:DNA Replication; D006801:Humans; D009421:Nervous System Malformations / Q000201:enzymology* / Q000235:genetics; D018809:Proliferating Cell Nuclear Antigen / Q000235:genetics / Q000378:metabolism; D055503:Protein Multimerization; D021381:Protein Transport; D016914:Ribonuclease H / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D018809:Proliferating Cell Nuclear Antigen; C091139:ribonuclease HII; D016914:Ribonuclease H