Abstract C018: Dissecting the Role of H3K4me3 in Transcriptional Activation Using a Dual-Inducible CRISPR-Based Epigenetic Effector Recruitment System

Abstract CRISPR-based epigenetic editing is a promising strategy for reprogramming gene expression by targeting chromatin-modifying enzymes to specific loci without altering the underlying DNA sequence. dCas9-mediated recruitment of the histone acetyltransferase P300 has been shown to activate gene expression through site-specific deposition of H3K27ac, a histone mark associated with active promoters and enhancers. The efficient transcriptional activation often requires the coordinated action of chromatin remodeling included changes in various histone modifications. Notably, the functional relationship between H3K27ac and H3K4me3, a histone mark enriched at transcription start sites, remains poorly defined in the context of synthetic epigenetic reprogramming. To address this, we developed a dual-inducible dCas9-based epigenetic editing platform that enables temporal and independent recruitment of different histone modifiers to defined genomic loci. This system integrates two orthogonal chemically inducible proximity (CIP) systems to separately control the deposition or removal of H3K27ac and H3K4me3. Using this platform, we interrogated the causal role of H3K4me3 in modulating transcriptional activation initiated by P300 recruitment. Our results demonstrated that H3K4me3 plays a critical role in shaping transcriptional kinetics and steady-state mRNA levels. Specifically, H3K4me3 influences both the early stages of transcriptional activation and the post-transcriptional stability of mRNA, suggesting a multifaceted role in regulating gene expression output. These findings indicate that H3K4me3 is not simply a passive marker of transcription but actively contributes to gene regulation in the whole life span of the mRNA transcript. In conclusion, our study provides a mechanistic framework for understanding the functional crosstalk between H3K4me3 and H3K27ac in transcriptional and posttranscriptional regulation. The dual-inducible CRISPR-based system offers a powerful and generalizable tool to investigate histone modification dynamics and may inform the rational design of next-generation epigenetic therapies. Citation Format: Chenwei Zhou, Fu-Sen Liang. Dissecting the Role of H3K4me3 in Transcriptional Activation Using a Dual-Inducible CRISPR-Based Epigenetic Effector Recruitment System [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2025 Oct 22-26; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2025;24(10 Suppl):Abstract nr C018.