The AREB1 Transcription Factor Influences Histone Acetylation To Regulate Drought Responses And Tolerance In Populus Trichocarpa

Perennial woody plants experience various environmental stresses in process of secondary growth.Among these stresses,drought is one of the key environmental limiting factors,which is a global threat to forestry production and natural ecosystem and seriously affects forest biomass and productivity.Transcription factors(TFs)and histone modifications play important roles in regulating plant adaptation to drought stress,but how they coordinate with each other to control the expression of drought-tolerant genes is unclear.Trees transport water absorbed from the soil to leaves and other organs by xylem.Xylem is a unique biological system for studying secondary growth and the adaptation to drought stress.Therefore,we study on a regulatory system involving coordinated regulation of H3K9 acetylation and AREB1 TF functions for activating many drought-responsive genes using the xylem of Populus trichocarpa.Genome-wide H3K9ac profiles and transcriptome changes in SDX tissues of P.trichocarpa under drought stress were generated using ChIP-seq and RNA-seq technique.Integrative analysis showed that increased H3K9ac level induces gene upregulation and decreased H3K9ac level induces gene downregulation.These results implied that histone modifications are involved in the regulation of gene expression in response to drought stress in P.trichocarpa.Analysis of motif enrichment motif searches revealed that the ABRE(ABA-Responsive Element)motif for the AREB1(ABA-Responsive Element Binding 1)-type protein was most significantly enriched within the H3K9ac-associated promoters and ABRE motifs likely mediate H3K9ac association and regulation of drought-response genes.In this study,we identified 76 drought-responsive TF DEGs that had the ABRE motif as well as differential H3K9ac levels in their promoters.We generated transgenic plants individually overexpressing 3 PtrNAC genes(PtrNAC006,PtrNAC007,PtrNAC120).Phynotype analysis showed that overexpression of PtrNAC genes resulted in smaller and more stem xylem vessel cells,and enhanced drought tolerance of these transgenic plants.It was suggested that PtrNAC genes affected secondary growth and reprogrammed cell differentiation for adaptation to drought stress.Both ChIP-qPCR and ChIP-seq data demonstrated that as the duration of drought treatment increased,H3K9ac enrichment increased progressively in all the identified ABRE motif regions of PtrNAC promoters.The results implied that ABRE motifs mediate H3K9ac association and regulation of PtrNAC genes in response to drought stress.RNA-seq and RT-qPCR data showed that AREB1 and histone acetyltransferase complexe genes were strongly induced by drought stress.In vitro Pull-down and in vivo BiFC analyses confirmed the forming of ternary protein complex.PtrGCN5-1 and PtrAREBl-2 downregulated transgenic plants,and PtrADA2b-3 CRISPR knockout mutants were also generated.ChIP-qPCR and RT-qPCR showed that reduced or deleted expression of PtrAREB1-2,PtrADA2b-3,or PtrGCN5-1 in P.trichocarpa decreases H3K9ac and RNA polymerase ?(Pol ?)enrichment on PtrNAC genes,expression of these NAC genes,and plant drought tolerance.In addition,our previous studies showed that AREB1 TF was able to bind the cis-elements of ABRE and upregulate the expression of PtrNAC genes.Taken together,the AREB1 TF interacts with the ADA2b-GCN5 histone acetyltransferase complex and recruits the complexes to PtrNAC006,007 and 120 genes through binding to ABRE motifs,resulting in enhanced H3K9ac and RNA(Pol II)enrichment for activating expression of the PtrNAC006,007 and 120 genes,thereby allowing P.trichocarpa for adaptation to drought stress.In conclusion,my desertation research confirmed the interaction of AREB1 TF and GCN5-ADA2b histone acetyltransferase complex,and their regulation of target genes.The research therefore uncovered the molecular mechanism of the coordinated histone acetylation and TF-mediated gene activation for drought response and tolerance in P.trichocarpa.The discovery has broad utility in improving drought tolerance of tree species and forest productivity.