Targeting histone acetylation to enhance somatic embryogenesis in <i>Quercus suber</i> L.

Somatic embryogenesis (SE) is an in vitro mass propagation system widely employed in plant breeding programs. However, its efficiency in many forest species remains limited due to their recalcitrance. Somatic embryogenesis relies on the induction of somatic cell reprogramming into embryogenic pathways, a process influenced by transcriptomic changes regulated, among other factors, by epigenetic modifications such as DNA methylation, histone methylation and histone acetylation. Despite its relevance, epigenetic regulation of SE in forest species is not well understood. In this study, we analyzed histone H4 acetylation during SE in cork oak (Quercus suber L.) and evaluated the effects of suberoylanilide hydroxamic acid (SAHA), a histone deacetylase (HDAC) inhibitor, scarcely used in plants, on the process. Histone H4 acetylation levels progressively increased after SE induction, correlating with enhanced histone acetyl transferase (HAT) enzymatic activity. The HAT gene QsHAM1-like was activated in developing somatic embryos, while HDAC genes QsHDA9, QsHDA19, QsHDA15 and QsHDA2 showed similar expression patterns among them, and opposite profiles to QsHAM1-like HAT gene, suggesting a coordinated interplay of HAT and HDAC activities in modulating global H4 acetylation during SE. SAHA treatment elevated histone H4 acetylation, promoted embryogenic mass proliferation, and induced the expression of QsSERK1-like, an early SE marker. While continuous SAHA exposure inhibited embryo differentiation, its removal restored embryo development, significantly increasing somatic embryo production. Inhibition of HAT activity by butyrolactone 3 decreased histone acetylation levels and reduced somatic embryo formation, providing further evidence that histone acetylation is essential for SE development. These findings highlight the critical role of histone acetylation in the SE of forest trees and propose transient treatments with epigenetic modulators like SAHA as a promising strategy to enhance somatic embryo production in recalcitrant forest species.