RRC ID 82555
Author Wu W, Zhu J, Nihira NT, Togashi Y, Goda A, Koike J, Yamaguchi K, Furukawa Y, Tomita T, Saeki Y, Johmura Y, Nakanishi M, Miyoshi Y, Ohta T.
Title Ribosomal S6 kinase (RSK) plays a critical role in DNA damage response via the phosphorylation of histone lysine demethylase KDM4B.
Journal Breast Cancer Res
Abstract BACKGROUND:Epigenetic dysregulation affecting oncogenic transcription and DNA damage response is a hallmark of cancer. The histone demethylase KDM4B, a factor regulating these processes, plays important roles in estrogen receptor-mediated transcription and DNA repair in breast cancer. However, how oncogenic phospho-signal transduction affects epigenetic regulation is not fully understood. Here we found that KDM4B phosphorylation by ribosomal S6 kinase (RSK), a downstream effector of the Ras/MAPK pathway, is critical for the function of KDM4B in response to DNA damage.
METHODS:KDM4B-knockout breast cancer cell lines were generated via CRISPR/Cas9-mediated gene editing. Re-expression of wild-type or phospho-site mutated KDM4B in knockout cells was performed by lentivirus-mediated gene transfer. Gene knockdown was achieved by RNA interference. DNA double-strand breaks (DSBs) were induced by ionizing radiation or laser-microirradiation. Protein accumulation at DSB sites was analyzed by immunofluorescence. KDM4B phosphorylation by RSK was assessed by in vitro and in vivo kinase assays. Gene and protein expression levels were analyzed by RT‒PCR and western blotting. The sensitivity of cells to ionizing radiation was examined by a clonogenic survival assay.
RESULTS:RSK phosphorylated KDM4B at Ser666, and inhibition of the phosphorylation by RSK depletion or RSK inhibitors abrogated KDM4B accumulation at the sites of DNA double-strand breaks (DSBs). DSB repair was significantly delayed in KDM4B-knockout cells or cells treated with RSK inhibitors. The replacement of endogenous KDM4B with the phosphomimetic mutant S666D restored KDM4B accumulation and DSB repair that had been inhibited by RSK inhibitors, suggesting a critical role for RSK at the specific serine residue of KDM4B in the effect of RSK inhibitors on DSB repair. As a consequence of these aberrant responses, inhibition of KDM4B phosphorylation increased the sensitivity of the cells to ionizing radiation.
CONCLUSIONS:Overall, the present study uncovered a novel function of RSK on the DNA damage response, which provides an additional role of its inhibitor in cancer therapy.
Volume 26(1)
Pages 146
Published 2024-10-21
DOI 10.1186/s13058-024-01901-x
PII 10.1186/s13058-024-01901-x
PMID 39434131
PMC PMC11492477
MeSH Breast Neoplasms* / genetics Breast Neoplasms* / metabolism Breast Neoplasms* / pathology Cell Line, Tumor DNA Breaks, Double-Stranded DNA Damage* DNA Repair* Female Gene Knockout Techniques Humans Jumonji Domain-Containing Histone Demethylases* / genetics Jumonji Domain-Containing Histone Demethylases* / metabolism Phosphorylation Ribosomal Protein S6 Kinases, 90-kDa / genetics Ribosomal Protein S6 Kinases, 90-kDa / metabolism
Resource
DNA material CSIV-TRE-RfA-UbC-KT (RDB12878) CS-RfA-ETBsd (RDB07917)