EIN2 mediates direct regulation of histone acetylation in the ethylene response

Abstract

Ethylene gas is essential for developmental processes and stress responses in plants. Although the membrane-bound protein EIN2 is critical for ethylene signaling, the mechanism by which the ethylene signal is transduced remains largely unknown. Here we show the levels of H3K14Ac and H3K23Ac are correlated with the levels of EIN2 protein and demonstrate EIN2 C terminus (EIN2-C) is sufficient to rescue the levels of H3K14/23Ac of ein2-5 at the target loci, using CRISPR/dCas9-EIN2-C. Chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) and ChIP-reChIP-seq analyses revealed that EIN2-C associates with histone partially through an interaction with EIN2 nuclear-associated protein1 (ENAP1), which preferentially binds to the genome regions that are associated with actively expressed genes both with and without ethylene treatments. Specifically, in the presence of ethylene, ENAP1-binding regions are more accessible upon the interaction with EIN2, and more EIN3 proteins bind to the loci where ENAP1 is enriched for a quick response. Together, these results reveal EIN2-C is the key factor regulating H3K14Ac and H3K23Ac in response to ethylene and uncover a unique mechanism by which ENAP1 interacts with chromatin, potentially preserving the open chromatin regions in the absence of ethylene; in the presence of ethylene, EIN2 interacts with ENAP1, elevating the levels of H3K14Ac and H3K23Ac, promoting more EIN3 binding to the targets shared with ENAP1 and resulting in a rapid transcriptional regulation.T he plant hormone ethylene is essential for a myriad of physiological and developmental processes. It is important in responses to stress, such as drought, cold, flooding, and pathogen infection (1, 2), and modulates stem cell division (3). The common aquatic ancestor of plants possessed the ethylene signaling pathway and the mechanism has been elucidated by analysis of Arabidopsis (4). Ethylene is perceived by a family of receptors bound to the membrane of the endoplasmic reticulum (ER) that are similar in sequence and structure to bacterial two-component histidine kinases (5-9). Each ethylene receptor has an N-terminal transmembrane domain, and the receptors form dimers that bind ethylene via a copper cofactor RAN1 (10, 11). Signaling from one of the receptors, ETR1, induces its physical association with the ER-localized protein RTE1 (12). The ethylene receptors function redundantly to negatively regulate ethylene responses (5) via a downstream Raf-like protein kinase called CTR1 (13,14).In the absence of ethylene, both the ethylene receptors and CTR1 are active, and CTR1 is associated with the ER membrane through a direct interaction with ETR1 (13). The CTR1 downstream factor, EIN2, is localized to the ER membrane, where it interacts with ETR1 (15). The protein stability of EIN2 is regulated by two F-box proteins, ETP1 and ETP2, which mediated its degradation by the ubiquitin-proteasome pathway (16). In the absence of ethylene, the CTR1-mediated phosphorylation at the C-terminal end of EIN2 (EIN2-C) leads to ...

Ethylene-triggered subcellular trafficking of CTR1 enhances the response to ethylene gas

“…Full- 25 length CTR1 also interacted with the EBFs in a BiFC assay ( Fig.5e). EIN2-CEND reconstituted YFP signals with its known nuclear interacting protein, EIN2 NUCLEAR-ASSOCIATED PROTEIN 1 (ENAP I) in the nucleus ( Supplementary Fig.5) 29 , showing that EIN2-CEND was present in the nucleus. Full-length CTR1 interacted with EBF1 and 2 regardless of ACC treatment ( Fig.5e and Supplementary Fig.6), which was consistent with constitutive nuclear localization of CTR1 in CaMV 35S overexpression lines ( Fig.1h).…”

CRISPR/Cas9‐Based Genome Editing and its Applications for Functional Genomic Analyses in Plants

“…Second, it has been fused with green fluorescent protein for the dynamic imaging of genomic loci . Third, the fusion of dCas9 to histone modifiers (e.g., methylase, demethylase, or acetyltransferase) or other proteins involved in DNA methylation has been used for targeted epigenetic regulation …”

REBELOTE, a regulator of floral determinacy in Arabidopsis thaliana, interacts with both nucleolar and nucleoplasmic proteins

“…It has been recently shown that VFP3/ENAP1 is involved in ethylene response. VFP3/ENAP1 interacts with EIN2, EIN3, SRT1 and SRT2 and regulates change in histone H3 acetylation levels and ethylene-dependant transcription [28][29][30]. Using a GFP transcriptional fusion protein, we investigated the subcellular localization of OBE1 and VFP3/ENAP1.…”