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Molecular Cancer

, 13:36

First Online: 24 February 2014Received: 30 July 2013Accepted: 21 January 2014DOI: 10.1186-1476-4598-13-36

Cite this article as: Cho, EA., Kim, EJ., Kwak, SJ. et al. Mol Cancer 2014 13: 36. doi:10.1186-1476-4598-13-36


BackgroundThe ataxia–telangiectasia mutated ATM protein kinase plays a central role in coordinating the cellular response to radiation-induced DNA damage. cAMP signaling regulates various cellular responses including metabolism and gene expression. This study aimed to investigate the mechanism through which cAMP signaling regulates ATM activation and cellular responses to ionizing radiation in lung cancer cells.

MethodsLung cancer cells were transfected with constitutively active stimulatory G protein GαsQL, and irradiated with γ-rays. The phosphorylation of ATM and protein phosphatase 2A was analyzed by western blotting, and apoptosis was assessed by western blotting, flow cytometry, and TUNNEL staining. The promoter activity of NF-κB was determined by dual luciferase reporter assay. BALB-c mice were treated with forskolin to assess the effect in the lung tissue.

ResultsTransient expression of GαsQL significantly inhibited radiation-induced ATM phosphorylation in H1299 human lung cancer cells. Treatment with okadaic acid or knock down of PP2A B56δ subunit abolished the inhibitory effect of Gαs on radiation-induced ATM phosphorylation. Expression of GαsQL increased phosphorylation of the B56δ and PP2A activity, and inhibition of PKA blocked Gαs-induced PP2A activation. GαsQL enhanced radiation-induced cleavage of caspase-3 and PARP and increased the number of early apoptotic cells. The radiation-induced apoptosis was increased by inhibition of NF-κB using PDTC or inhibition of ATM using KU55933 or siRNA against ATM. Pretreatment of BALB-c mice with forskolin stimulated phosphorylation of PP2A B56δ, inhibited the activation of ATM and NF-κB, and augmented radiation-induced apoptosis in the lung tissue. GαsQL expression decreased the nuclear levels of the p50 and p65 subunits and NF-κB-dependent activity after γ-ray irradiation in H1299 cells. Pretreatment with prostaglandin E2 or isoproterenol increased B56δ phosphorylation, decreased radiation-induced ATM phosphorylation and increased apoptosis.

ConclusionscAMP signaling inhibits radiation-induced ATM activation by PKA-dependent activation of PP2A, and this signaling mechanism augments radiation-induced apoptosis by reducing ATM-dependent activation of NF-κB in lung cancer cells.

KeywordscAMP signaling ATM Protein phosphatase 2A Apoptosis NF-κB Lung cancer AbbreviationsATMAtaxia–telangiectasia mutated

CREBcAMP response element-binding protein

GαsStimulatory α subunit of G protein

GαsQLConstitutively active mutant long form of the α subunit of stimulatory heterotrimeric GTP binding protein

PKAcAMP-dependent protein kinase

PP2AProtein phosphatase 2A

PARPPoly ADP adenosine diphosphate-ribose polymerase

qPCRQuantitative polymerase chain reaction

siRNASmall interfering RNA

TUNELTerminal uridine nucleotide end-labeling

Electronic supplementary materialThe online version of this article doi:10.1186-1476-4598-13-36 contains supplementary material, which is available to authorized users.

Eun-Ah Cho, Eui-Jun Kim contributed equally to this work.

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Autor: Eun-Ah Cho - Eui-Jun Kim - Sahng-June Kwak - Yong-Sung Juhnn


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