Inhibition of HDAC7 reprograms the histone H3.3 landscape to induce heterochromatin spreading and DNA replication...

Class IIa histone deacetylases (HDACs) have minimal HDAC activity but may function as multiprotein interaction hubs. Here, we demonstrated the expression of histone deacetylase 7 (HDAC7), a class IIa HDAC family member, in glioblastoma tumor tissue from 84 patients, patient-derived glioma stem cells from six patients, and pediatric diffuse pontine glioma cells from three patients. HDAC7 binds to histone H3.3 and interacts with H3.3 and HIRA on chromatin. Targeted downregulation of HDAC7 expression with a subtype-specific siRNA inhibits the interaction of H3.3 with HIRA, increasing the association of H3.3 with DAXX and histone lysine trimethylation at lysine 9 (H3K9me3), leading to H3.3 deposition on H3K9me3⁺/4',6-diamidino-2-phenylindole⁺ heterochromatin nuclear foci. HDAC7 inhibition triggers H3K9me3⁺ heterochromatin spreading, increased H3K9me3 binding in the cancer genome, and significant alterations in gene expression. Using single-molecule DNA fiber analysis, we showed that HDAC7 inhibition resulted in a significant increase in replication fork speed without affecting fork symmetry, leading to replication stress, phosphorylation of RPA2, and reduced 5-ethynyl-2'-deoxyuridine incorporation. HDAC7 depletion also reduces BRCA2 expression and increases cancer cell sensitivity to DNA-damaging agents. These findings reveal the role of HDAC7 in the euchromatic H3.3 chaperone network and the impact of HDAC7 depletion on chromatin dynamics, epigenetic restriction, and DNA damage in cancer cells.