title: DNA damage response (DDR) induced by topoisomerase II poisons requires nuclear function of the small GTPase Rac.
abstract: Here, we investigated the influence of Rac family small GTPases on mechanisms of the DNA damage response (DDR) stimulated by topoisomerase II poisons. To this end, we examined the influence of the Rac-specific small molecule inhibitor EHT1864 on Ser139 phosphorylation of histone H2AX, a widely used marker of the DDR triggered by DNA double-strand breaks. EHT1864 attenuated the doxorubicin-stimulated DDR in a subset of cell lines tested, including HepG2 hepatoma cells. EHT1864 reduced the level of DNA strand breaks and increased viability following treatment of HepG2 cells with topo II poisons. Protection by EHT1864 was observed in both p53 wildtype (HepG2) and p53 deficient (Hep3B) human hepatoma cells and, furthermore, remained unaffected upon pharmacological inhibition of p53 in HepG2. Apparently, the impact of Rac on the DDR is independent of p53. Protection from doxorubicin-induced DNA damage by EHT1864 comprises both S and G2 phase cells. The inhibitory effect of EHT1864 on doxorubicin-stimulated DDR was mimicked by pharmacological inhibition of various protein kinases, including JNK, ERK, PI3K, PAK and CK1. EHT1864 and protein kinase inhibitors also attenuated the formation of the topo II-DNA cleavable complex. Moreover, EHT1864 mitigated the constitutive phosphorylation of topoisomerase IIα at positions S1106, S1213 and S1247. Doxorubicin transport, nuclear import/export of topoisomerase II and Hsp90-related mechanisms are likely not of relevance for doxorubicin-stimulated DDR impaired by EHT1864. We suggest that multiple kinase-dependent but p53- and heat shock protein-independent Rac-regulated nuclear mechanisms are required for activation of the DDR following treatment with topo II poisons.
authors.affiliation: Institute of Toxicology, Heinrich-Heine-University Düsseldorf, Moorenstrasse 5, D-40225 Düsseldorf, Germany.
authors.forename: Friedrich
authors.initials: F
authors.lastname: Wartlick
authors.affiliation: Institute of Toxicology, Heinrich-Heine-University Düsseldorf, Moorenstrasse 5, D-40225 Düsseldorf, Germany.
authors.forename: Anita
authors.initials: A
authors.lastname: Bopp
authors.affiliation: Institute of Toxicology, Heinrich-Heine-University Düsseldorf, Moorenstrasse 5, D-40225 Düsseldorf, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Henninger
authors.affiliation: Institute of Toxicology, Heinrich-Heine-University Düsseldorf, Moorenstrasse 5, D-40225 Düsseldorf, Germany. Electronic address: fritz@uni-duesseldorf.de.
authors.forename: Gerhard
authors.initials: G
authors.lastname: Fritz
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D000818:Animals; D000951:Antigens, Neoplasm / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000201:enzymology*; D019610:Cytoprotection / Q000187:drug effects; D004249:DNA Damage; D004250:DNA Topoisomerases, Type II / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D004317:Doxorubicin / Q000494:pharmacology; D016195:G2 Phase / Q000187:drug effects; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011753:Pyrones / Q000494:pharmacology; D011804:Quinolines / Q000494:pharmacology; D051381:Rats; D016196:S Phase / Q000187:drug effects; D015398:Signal Transduction / Q000187:drug effects; D059005:Topoisomerase II Inhibitors / Q000494:pharmacology*; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D020744:rac GTP-Binding Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*
chemical_list: D000951:Antigens, Neoplasm; D004268:DNA-Binding Proteins; C506907:EHT 1864; D018841:HSP90 Heat-Shock Proteins; D047428:Protein Kinase Inhibitors; D011753:Pyrones; D011804:Quinolines; D059005:Topoisomerase II Inhibitors; D016159:Tumor Suppressor Protein p53; D004317:Doxorubicin; D020744:rac GTP-Binding Proteins; D004250:DNA Topoisomerases, Type II
keywords: DNA damage response; Rac family small GTPase; Topoisomerase II poisons