Bru-seq analysis of nascent transcription in IDH1-R132H mutant versus wild-type mouse glioma neurospheres

Using a genetically engineered mouse glioma model (Sleeping Beauty transposon system; NRAS(G12V), shRNA-mediated Trp53 and Atrx knockdown, with or without mutant IDH1-R132H) and derived neurosphere (NS) cultures, we asked whether IDH1-R132H alters nascent transcription of DNA damage response (DDR) genes. Nascent RNA was metabolically labeled with 5-bromouridine (Bru) and Bru-labeled transcripts were immunopurified and sequenced (Bru-seq) to measure genome-wide nascent transcription rates at baseline (untreated), independent of steady-state mRNA abundance.

Bru-seq identified increased nascent transcription of DDR genes, including Atm and Rad50, in IDH1-R132H-mutant (mIDH1) NS relative to wild-type IDH1 (wt-IDH1) NS, consistent with the epigenetic up-regulation of ATM signaling and enhanced DDR reported for this model.