HUS1 regulates in vivo responses to genotoxic chemotherapies.
Cells are under constant attack from genotoxins and rely on a multifaceted DNA damage response (DDR) network to maintain genomic integrity. Central to the DDR are the ATM and ATR kinases, which respond primarily to double-strand DNA breaks (DSBs) and replication stress, respectively. Optimal ATR signaling requires the RAD9A-RAD1-HUS1 (9-1-1) complex, a toroidal clamp that is loaded at damage sites and scaffolds signaling and repair factors. Whereas complete ATR pathway inactivation causes embryonic lethality, partial Hus1 impairment has been accomplished in adult mice using hypomorphic (Hus1(neo)) and null (Hus1(Δ1)) Hus1 alleles, and here we use this system to define the tissue- and cell type-specific actions of the HUS1-mediated DDR in vivo. Hus1(neo/Δ1) mice showed hypersensitivity to agents that cause replication stress, including the crosslinking agent mitomycin C (MMC) and the replication inhibitor hydroxyurea, but not the DSB inducer ionizing radiation. Analysis of tissue morphology, genomic instability, cell proliferation and apoptosis revealed that MMC treatment caused severe damage in highly replicating tissues of mice with partial Hus1 inactivation. The role of the 9-1-1 complex in responding to MMC was partially ATR-independent, as a HUS1 mutant that was proficient for ATR-induced checkpoint kinase 1 phosphorylation nevertheless conferred MMC hypersensitivity. To assess the interplay between the ATM and ATR pathways in responding to replication stress in vivo, we used Hus1/Atm double mutant mice. Whereas Hus1(neo/neo) and Atm(-/-) single mutant mice survived low-dose MMC similar to wild-type controls, Hus1(neo/neo)Atm(-/-) double mutants showed striking MMC hypersensitivity, consistent with a model in which MMC exposure in the context of Hus1 dysfunction results in DSBs to which the ATM pathway normally responds. This improved understanding of the inter-dependency between two major DDR mechanisms during the response to a conventional chemotherapeutic illustrates how inhibition of checkpoint factors such as HUS1 may be effective for the treatment of ATM-deficient and other cancers.
Department of Biomedical Sciences, Cornell University, Ithaca, NY, USA.
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Balmus
Department of Biomedical Sciences, Cornell University, Ithaca, NY, USA.
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Lim
Department of Biomedical Sciences, Cornell University, Ithaca, NY, USA.
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Oswald
Department of Biomedical Sciences, Cornell University, Ithaca, NY, USA.|Department of Clinical Sciences, Cornell University, Ithaca, NY, USA.
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Hume
Department of Biomedical Sciences, Cornell University, Ithaca, NY, USA.
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Cassano
Department of Biomedical Sciences, Cornell University, Ithaca, NY, USA.
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Pierre
Department of Biomedical Sciences, Cornell University, Ithaca, NY, USA.
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Hill
Department of Microbiology and Immunology, Cornell University, Ithaca, NY, USA.
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Huang
Department of Microbiology and Immunology, Cornell University, Ithaca, NY, USA.
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http://orcid.org/0000-0003-4968-3415
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August
Department of Population Medicine and Diagnostic Sciences, Cornell University, Ithaca, NY, USA.
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Stokol
Department of Biomedical Sciences, Cornell University, Ithaca, NY, USA.
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Southard
Department of Biomedical Sciences, Cornell University, Ithaca, NY, USA.
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Weiss
D000818:Animals; D064007:Ataxia Telangiectasia Mutated Proteins / Q000235:genetics / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D004249:DNA Damage; D051379:Mice; D009152:Mutagenicity Tests; D009153:Mutagens / Q000494:pharmacology*; D015398:Signal Transduction
D018797:Cell Cycle Proteins; C509939:Hus1 protein, mouse; D009153:Mutagens; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; C576325:Atm protein, mouse