ZFP36L2 orchestrates stress-adaptive plasticity in intestinal regeneration and colorectal cancer metastasis (ActDseq)

Organoids expressing a doxycycline-inducible short hairpin RNA targeting ZFP36L2 (shZFP36L2) or a non-targeting control (shCtrl) were cultured for 5 days with 2 μg/mL doxycycline, then treated with or without 5 μg/mL Actinomycin D (ActD) for 5 hours to block de novo transcription.

Total RNA was extracted using the RNeasy Mini Kit (Qiagen), and strand-specific mRNA libraries were prepared using the TruSeq Stranded mRNA Kit and sequenced on the Illumina NovaSeq 6000 (PE100). Decay rates were quantified by comparing transcript abundance between ActD-treated and untreated conditions within each knockdown condition; ZFP36L2-dependent decay was calculated as the differential decay rate between shZFP36L2 and shCtrl.