Ethylene regulates many aspects of the plant life cycle, including seed germination, root initiation, flower development, fruit ripening, senescence, and responses to biotic and abiotic stresses. It thus plays a key role in responses to the environment that have a direct bearing on a plant's fitness for adaptation and reproduction. In recent years, there have been major advances in our understanding of the molecular mechanisms regulating ethylene synthesis and action. Screening for mutants of the triple response phenotype of etiolated Arabidopsis seedlings, together with map-based cloning and candidate gene characterization of natural mutants from other plant species, has led to the identification of many new genes for ethylene biosynthesis, signal transduction, and response pathways. The simple chemical nature of ethylene contrasts with its regulatory complexity. This is illustrated by the multiplicity of genes encoding the key ethylene biosynthesis enzymes 1-aminocyclopropane-1-carboxylic acid (ACC) synthase and ACC oxidase, multiple ethylene receptors and signal transduction components, and the complexity of regulatory steps involving signalling relays and control of mRNA and protein synthesis and turnover. In addition, there are extensive interactions with other hormones. This review integrates knowledge from the model plant Arabidopsis and other plant species and focuses on key aspects of recent research on regulatory networks controlling ethylene synthesis and its role in flower development and fruit ripening.

1-Aminocyclopropane-1-carboxylic acid synthase (ACS) is one of the key regulatory enzymes involved in the synthesis of the hormone ethylene and is encoded by a multigene family containing at least eight members in tomato (Lycopersicon esculentum). Increased ethylene production accompanies ripening in tomato, and this coincides with a change in the regulation of ethylene synthesis from auto-inhibitory to autostimulatory. The signaling pathways that operate to bring about this transition from so-called system-1 to system-2 ethylene production are unknown, and we have begun to address these by investigating the regulation of ACS expression during ripening. Transcripts corresponding to four ACS genes,LEACS1A, LEACS2, LEACS4, and LEACS6, were detected in tomato fruit, and expression analysis using the ripening inhibitor(rin) mutant in combination with ethylene treatments and the Never-ripe (Nr) mutant has demonstrated that each is regulated in a unique way. A proposed model suggests that system-1 ethylene is regulated by the expression ofLEACS1A and LEACS6. In fruit a transition period occurs in which the RIN gene plays a pivotal role leading to increased expression of LEACS1A and induction of LEACS4. System-2 ethylene synthesis is subsequently initiated and maintained by ethylene-dependent induction ofLEACS2.

Fleshy fruits using ethylene to regulate ripening have developed multiple times in the history of angiosperms, presenting a clear case of convergent evolution whose molecular basis remains largely unknown. Analysis of the fruitENCODE data consisting of 361 transcriptome, 71 accessible chromatin, 147 histone and 45 DNA methylation profiles reveals three types of transcriptional feedback circuits controlling ethylene-dependent fruit ripening. These circuits are evolved from senescence or floral organ identity pathways in the ancestral angiosperms either by neofunctionalisation or repurposing pre-existing genes. The epigenome, H3K27me3 in particular, has played a conserved role in restricting ripening genes and their orthologues in dry and ethylene-independent fleshy fruits. Our findings suggest that evolution of ripening is constrained by limited hormone molecules and genetic and epigenetic materials, and whole-genome duplications have provided opportunities for plants to successfully circumvent these limitations.

There are at least five lipoxygenases (TomloxA, TomloxB, TomloxC, TomloxD, and TomloxE) present in tomato (Lycopersicon esculentum Mill.) fruit, but their role in generation of fruit flavor volatiles has been unclear. To assess the physiological role of TomloxC in the generation of volatile C6 aldehyde and alcohol flavor compounds, we produced transgenic tomato plants with greatly reduced TomloxC using sense and antisense constructs under control of the cauliflower mosaic virus 35S promoter. The expression level of the TomloxC mRNA in some transgenic plants was selectively reduced by gene silencing or antisense inhibition to between 1% and 5% of the wild-type controls, but the expression levels of mRNAs for the four other isoforms were unaffected. The specific depletion of TomloxC in transgenic tomatoes led to a marked reduction in the levels of known flavor volatiles, including hexanal, hexenal, and hexenol, to as little as 1.5% of those of wild-type controls following maceration of ripening fruit. Addition of linoleic or linolenic acid to fruit homogenates significantly increased the levels of flavor volatiles, but the increase with the TomloxC-depleted transgenic fruit extracts was much lower than with the wild-type control. Confocal imaging of tobacco (Nicotiana tabacum) leaf cells expressing a TomloxC-GFP fusion confirmed a chloroplast localization of the protein. Together, these results suggest that TomloxC is a chloroplast-targeted lipoxygenase isoform that can use both linoleic and linolenic acids as substrates to generate volatile C6 flavor compounds. The roles of the other lipoxygenase isoforms are discussed.

Ethylene response factors (ERFs) are downstream components of ethylene-signaling pathways known to play critical roles in ethylene-controlled climacteric fruit ripening, yet little is known about the molecular mechanism underlying their mode of action. Here, we demonstrate that SlERF.F12, a member of the ERF.F subfamily containing Ethylene-responsive element-binding factor-associated Amphiphilic Repression (EAR) motifs, negatively regulates the onset of tomato (Solanum lycopersicum) fruit ripening by recruiting the co-repressor TOPLESS 2 (TPL2) and the histone deacetylases (HDAs) HDA1/HDA3 to repress the transcription of ripening-related genes. The SlERF.F12-mediated transcriptional repression of key ripening-related genes 1-AMINO-CYCLOPROPANE-1-CARBOXYLATE SYNTHASE 2 (ACS2), ACS4, POLYGALACTURONASE 2a, and PECTATE LYASE is dependent on the presence of its C-terminal EAR motif. We show that SlERF.F12 interacts with the co-repressor TPL2 via the C-terminal EAR motif and recruits HDAs SlHDA1 and SlHDA3 to form a tripartite complex in vivo that actively represses transcription of ripening genes by decreasing the level of the permissive histone acetylation marks H3K9Ac and H3K27Ac at their promoter regions. These findings provide new insights into the ripening regulatory network and uncover a direct link between repressor ERFs and histone modifiers in modulating the transition to ripening of climacteric fruit.