title: The leukemia-associated Rho guanine nucleotide exchange factor LARG is required for efficient replication stress signaling.
abstract: We previously identified and characterized TELO2 as a human protein that facilitates efficient DNA damage response (DDR) signaling. A subsequent yeast 2-hybrid screen identified LARG; Leukemia-Associated Rho Guanine Nucleotide Exchange Factor (also known as Arhgef12), as a potential novel TELO2 interactor. LARG was previously shown to interact with Pericentrin (PCNT), which, like TELO2, is required for efficient replication stress signaling. Here we confirm interactions between LARG, TELO2 and PCNT and show that a sub-set of LARG co-localizes with PCNT at the centrosome. LARG-deficient cells exhibit replication stress signaling defects as evidenced by; supernumerary centrosomes, reduced replication stress-induced γH2AX and RPA nuclear foci formation, and reduced activation of the replication stress signaling effector kinase Chk1 in response to hydroxyurea. As such, LARG-deficient cells are sensitive to replication stress-inducing agents such as hydroxyurea and mitomycin C. Conversely we also show that depletion of TELO2 and the replication stress signaling kinase ATR leads to RhoA signaling defects. These data therefore reveal a level of crosstalk between the RhoA and DDR signaling pathways. Given that mutations in both ATR and PCNT can give rise to the related primordial dwarfism disorders of Seckel Syndrome and Microcephalic osteodysplastic primordial dwarfism type II (MOPDII) respectively, which both exhibit defects in ATR-dependent checkpoint signaling, these data also raise the possibility that mutations in LARG or disruption to RhoA signaling may be contributory factors to the etiology of a sub-set of primordial dwarfism disorders.
authors.affiliation: a Genome Stability Group ; Department of Oncology ; Academic Unit of Molecular Oncology ; University of Sheffield Medical School ; Sheffield , UK.
authors.forename: Ryan D
authors.initials: RD
authors.lastname: Beveridge
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Staples
authors.forename: Abhijit A
authors.initials: AA
authors.lastname: Patil
authors.forename: Katie N
authors.initials: KN
authors.lastname: Myers
authors.forename: Sarah
authors.initials: S
authors.lastname: Maslen
authors.forename: J Mark
authors.initials: JM
authors.lastname: Skehel
authors.forename: Simon J
authors.initials: SJ
authors.lastname: Boulton
authors.forename: Spencer J
authors.initials: SJ
authors.lastname: Collis
mesh_terms: D000941:Antigens / Q000378:metabolism; D064007:Ataxia Telangiectasia Mutated Proteins / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D018385:Centrosome / Q000378:metabolism; D000071877:Checkpoint Kinase 1; D004260:DNA Repair; D004261:DNA Replication / Q000187:drug effects; D045325:HCT116 Cells; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D006918:Hydroxyurea / Q000494:pharmacology; D016685:Mitomycin / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011494:Protein Kinases / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D064067:Rho Guanine Nucleotide Exchange Factors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D034501:Telomere-Binding Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000378:metabolism
chemical_list: C577222:ARHGEF12 protein, human; D000941:Antigens; D034741:RNA, Small Interfering; D064067:Rho Guanine Nucleotide Exchange Factors; D034501:Telomere-Binding Proteins; C086178:pericentrin; D016685:Mitomycin; D011494:Protein Kinases; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; D020742:rhoA GTP-Binding Protein; D006918:Hydroxyurea
keywords: ATR signalling; DDR, DNA damage response; DNA damage response; HU, hydroxyurea; PCNT, pericentrin; RhoGEF; centrosome; replication stress