Project description:We report that ethylene regulate the ENAP1 binding. We perform a dual cross-linking procedure using the additional cross-linker ethylene glycol bis-succinimidylsuccinate (EGS; Thermo Scientific) along with formaldehyde according to (Pedmale et al. 2016) for Chip-sequencing (ChIPseq) of ENAP1, as well as ENAP1 ChIP- EIN2 reChIPseq (ChIP-reChIPseq), which were used chromatins isolated from 3-day old etiolated Col-0 seedlings treated with ethylene or air gas. Results show that ENAP1 is involved in the EIN2 dependent ethylene response.
Project description:We report that ethylene reduce the ENAP1 binding. We perform Chip-sequencing of ENAP1 using chromatins isolated from 3-day old etiolated ein3eil1 seedlings treated with ethylene or air gas. Results show that ethylene reduce ENAP1 binding in TSS region of EIN3 targeted genes that are regulated at the transcriptional level by ethylene.
Project description:We report that ethylene reduce the ENAP1 binding. We perform Chip-sequencing of ENAP1 using chromatins isolated from 3-day old etiolated Col-0/ein2/ein3eil1 seedlings treated with ethylene or air gas. Results show that ethylene reduce ENAP1 binding in TSS region of EIN3 targeted genes that are regulated at the transcriptional level by ethylene.
Project description:A complex interplay between ethylene, ETP1/ETP2 F-box proteins, and degradation of EIN2 is essential for triggering ethylene responses in plants. The gaseous plant hormone ethylene can trigger myriad physiological and morphological responses in plants. While many ethylene signaling pathway components have been identified and characterized, little is known about the function of the integral membrane protein EIN2, a central regulator of all ethylene responses. Here, we demonstrate that Arabidopsis thaliana EIN2 is a protein with a short half-life that undergoes rapid proteasome-mediated protein turnover. Moreover, EIN2 protein accumulation is positively regulated by ethylene. We identified two F-box proteins, EIN2 TARGETING PROTEIN and 2 (ETP1 and ETP2), that interact with the EIN2 carboxyl-terminal domain (CEND), which is highly conserved and sufficient to activate most ethylene responses. Overexpression of ETP1 or ETP2 disrupts EIN2 protein accumulation, and these plants manifest a strong ethylene insensitive phenotype. Furthermore, knocking down the levels of both ETP1 and ETP2 mRNAs using an artificial microRNA (amiRNA) leads to accumulation of EIN2 protein, resulting in plants that display constitutive ethylene response phenotypes. Finally, ethylene down-regulates ETP1 and ETP2 proteins, impairing their ability to interact with EIN2. Thus, these studies reveal that a complex interplay between ethylene, the regulation of ETP1/ETP2 F-box proteins, and subsequent targeting and degradation of EIN2 is essential for triggering ethylene responses in plants. Keywords: ethylene treatment, genetic modification
Project description:A complex interplay between ethylene, ETP1/ETP2 F-box proteins, and degradation of EIN2 is essential for triggering ethylene responses in plants. The gaseous plant hormone ethylene can trigger myriad physiological and morphological responses in plants. While many ethylene signaling pathway components have been identified and characterized, little is known about the function of the integral membrane protein EIN2, a central regulator of all ethylene responses. Here, we demonstrate that Arabidopsis thaliana EIN2 is a protein with a short half-life that undergoes rapid proteasome-mediated protein turnover. Moreover, EIN2 protein accumulation is positively regulated by ethylene. We identified two F-box proteins, EIN2 TARGETING PROTEIN and 2 (ETP1 and ETP2), that interact with the EIN2 carboxyl-terminal domain (CEND), which is highly conserved and sufficient to activate most ethylene responses. Overexpression of ETP1 or ETP2 disrupts EIN2 protein accumulation, and these plants manifest a strong ethylene insensitive phenotype. Furthermore, knocking down the levels of both ETP1 and ETP2 mRNAs using an artificial microRNA (amiRNA) leads to accumulation of EIN2 protein, resulting in plants that display constitutive ethylene response phenotypes. Finally, ethylene down-regulates ETP1 and ETP2 proteins, impairing their ability to interact with EIN2. Thus, these studies reveal that a complex interplay between ethylene, the regulation of ETP1/ETP2 F-box proteins, and subsequent targeting and degradation of EIN2 is essential for triggering ethylene responses in plants. Experiment Overall Design: Six samples were analyzed. There were three treatments with two biological replicates each. The treatments are as follows: 8 week old Col-0 plants (air control), 8 week old Col-0 plants treated for 24 hours with ethylene gas (10 ppm), and artificial microRNA knockdown mutants, amiR-ETP1/2.
Project description:Ethylene gas is essential for many developmental processes and stress responses in plants. ETHYLENE INSENSITIVE2 (EIN2), an NRAMP-homologous integral membrane protein, plays an essential role in ethylene signaling but its function remains enigmatic. Here we report that phosphorylation-regulated proteolytic processing of EIN2 triggers its endoplasmic reticulum (ER)-nucleus translocation, which is essential for hormone signaling and response in Arabidopsis. Without ethylene, or in hormone receptors mutants, ER-tethered EIN2 shows CTR1 kinase-dependent phosphorylation. Ethylene exposure triggers dephosphorylation and proteolytic cleavage, resulting in rapid nuclear translocation of a carboxyl-terminal EIN2 fragment (C’). Plants containing mutations that mimic EIN2 dephosphorylation, or inactivate CTR1, show constitutive cleavage and nuclear localization of EIN2-C’, and EIN3/EIL1-dependent activation of ethylene responses. These findings uncover a mechanism of subcellular communication whereby ethylene gas stimulates rapid phosphorylation-dependent cleavage and nuclear movement of the EIN2-C’ peptide, thus linking hormone perception and signaling components located in the ER with nuclear-localized transcriptional regulators.
Project description:Purpose: Next-generation sequencing (NGS) has been used to study the gene expression in different samples under air and ethylene treatment. The goal of this study is to uncover how ENAP1 and H3K23Ac dynamically coordinate with EIN3 to regulate gene expression in response to ethylene.
Project description:Purpose: Next-generation sequencing (NGS) has been used to study the gene expression in different samples under air and ethylene treatment. The goal of this study is to uncover how ENAP1 and H3K23Ac dynamically coordinate with EIN3 to regulate gene expression in response to ethylene.
