ABSTRACT
We studied Arabidopsis HYPOXIA-RESPONSIVE MODULATOR 1 (HRM1), which belongs to a group of core hypoxia-responsive genes that are conserved among plant species across great evolutionary distance. The hrm1 mutants had lower survival rates and showed more damage than the wild-type (WT) plants under hypoxic stress. Promoter analyses showed that HRM1 is regulated by EIN3 and RAP2.2 during hypoxia. Fluorescence tracing and immunogold labeling assays showed that HRM1 protein was enriched in mitochondria. Co-immunoprecipitation coupled with mass spectrometry and bimolecular fluorescence complementation assays showed that HRM1 associates with the complex-I in mitochondria. Compared with the WT plants, metabolic activities related to the mitochondrial electron transport chain (mETC) were higher in hrm1 mutants during hypoxia. Loss of HRM1 caused de-repression of mETC complex I, II, and IV activities and higher basal and maximum respiration rates under hypoxia. Our results showed that through association with complex-I, HRM1 attenuates mETC activity and modulates the respiratory chain under low oxygen. Compared with the regulatory system in mammalian, adjustment of mitochondrial respiration to low oxygen helps plants decrease reactive oxygen species production and is also critical for the submergence survival.
Subject(s)
Arabidopsis Proteins , Arabidopsis , Animals , Arabidopsis/genetics , Arabidopsis/metabolism , Electron Transport , Hypoxia , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Oxygen/metabolism , MammalsABSTRACT
Ethylene is an essential hormone in plants that is involved in low-oxygen and reoxygenation responses. As a key transcription factor in ethylene signaling, ETHYLENE INSENSITIVE3 (EIN3) activates targets that trigger various responses. However, most of these targets are still poorly characterized. Through analyses of our microarray data and the published Arabidopsis (Arabidopsis thaliana) EIN3 chromatin immunoprecipitation sequencing data set, we inferred the putative targets of EIN3 during anoxia-reoxygenation. Among them, GDH2, which encodes one subunit of glutamate dehydrogenase (GDH), was chosen for further studies for its role in tricarboxylic acid cycle replenishment. We demonstrated that both GDH1 and GDH2 are induced during anoxia and reoxygenation and that this induction is mediated via ethylene signaling. In addition, the results of enzymatic assays showed that the level of GDH during anoxia-reoxygenation decreased in the ethylene-insensitive mutants ein2-5 and ein3eil1 Global metabolite analysis indicated that the deamination activity of GDH might regenerate 2-oxoglutarate, which is a cosubstrate that facilitates the breakdown of alanine by alanine aminotransferase when reoxygenation occurs. Moreover, ineffective tricarboxylic acid cycle replenishment, disturbed carbohydrate metabolism, reduced phytosterol biosynthesis, and delayed energy regeneration were found in gdh1gdh2 and ethylene mutants during reoxygenation. Taken together, these data illustrate the essential role of EIN3-regulated GDH activity in metabolic adjustment during anoxia-reoxygenation.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/enzymology , Arabidopsis/metabolism , Ethylenes/pharmacology , Glutamate Dehydrogenase/metabolism , Oxygen/metabolism , Anaerobiosis/drug effects , Arabidopsis/drug effects , Arabidopsis/genetics , Carbohydrate Metabolism/drug effects , DNA-Binding Proteins , Energy Metabolism/drug effects , Gene Expression Regulation, Plant/drug effects , Metabolome/drug effects , Models, Biological , Nuclear Proteins/metabolism , Phenotype , Phytosterols/biosynthesis , Protein Stability/drug effects , Signal Transduction/drug effects , Transcription Factors/metabolismABSTRACT
Ethylene is known to play an essential role in mediating hypoxic responses in plants. Here, we show that in addition to regulating hypoxic responses, ethylene also regulates cellular responses in the reoxygenation stage after anoxic treatment in Arabidopsis. We found that expression of several ethylene biosynthetic genes and ethylene-responsive factors, including ERF1 and ERF2, was induced during reoxygenation. Compared with the wild type, two ethylene-insensitive mutants (ein2-5 and ein3eil1) were more sensitive to reoxygenation and displayed damaged phenotypes during reoxygenation. To characterize the role of ethylene, we applied microarray analysis to Col-0, ein2-5 and ein3eil1 under reoxygenation conditions. Our results showed that gene transcripts involved in reactive oxygen species (ROS) detoxification, dehydration response and metabolic processes were regulated during reoxygenation. Moreover, ethylene signalling may participate in regulating these responses and maintaining the homeostasis of different phytohormones. Our work presents evidence that ethylene has distinct functions in recovery after anoxia and provides insight into the reoxygenation signalling network.
