Ethylene is a gaseous phytohormone involved in multiple aspects of plant growth, development, senescence, and stress response. Seedlings that are germinated in the dark in the presence of ethylene undergo specific phenotypic changes known as the triple response. The three elements of this response are the radial expansion and growth inhibition of hypocotyls and roots and an exaggeration of the apical hook curvature. At the molecular level, the developmental effects of ethylene are accompanied by significant changes in gene expression at both transcriptional and post-transcriptional levels. While transcriptional regulation is well established as a critical process in response to ethylene, little is known about the role of ethylene-triggered gene-specific regulation of translation. Through ribosomal footprinting, our group uncovered a critical molecular mechanism that links ethylene perception to the activation of a novel gene-specific translational control mechanism. Characterization of one of the targets of this translational regulation, EBF2, indicated that the signaling molecule EIN2 and the nonsense-mediated decay proteins UPFs play a central role in this ethylene-induced translational response, setting a new paradigm of gene-specific translational control. We aim to test the role of additional candidate genes whose translational efficiency is affected by ethylene. I am characterizing T-DNA knockouts corresponding to ethylene-responsive translational targets and studying their growth dynamics through a growth response kinetic assay. This test relies on an infrared live imaging system to monitor subtle changes in the rates of elongation in hypocotyls and roots of dark-grown seedlings transiently exposed to the ethylene gas. In parallel, I am also exploring changes in the hypocotyl and root elongation in previously characterized ethylene- and auxin-insensitive mutants. Auxin is another vital plant hormone that controls numerous processed in plant’s life cycle, from embryo development to fruit ripening. Remarkably, auxin biosynthesis, transport, and signaling are known to be interconnected with the ethylene biosynthesis and signaling pathway.
Thus, mutant plants with defects in auxin also show phenotypic deviations in their response to ethylene. I have tested a set of previously characterized auxin mutants regarding their dynamic responses to ethylene to determine which stages of the ethylene response and recovery are compromised. My data indicate that auxin is required for the early stage of the ethylene response (aka the fast response), that is thought to be independent of transcription. However, transcription independence of the fast ethylene response has not been fully proven, as no loss of function mutants are available for all of the candidate transcriptional master regulators of the ethylene signaling pathway. ETHYLENE INSENSITIVE3/ETHYLENE INSENSITIVE3-LIKE (EIN3/EIL1) are the only well-characterized transcriptional master regulators of the ethylene response. The functions of their putative orthologs EIL2, EIL3, EIL4, and EIL5 are unknown. We aim to generate CRISPR/Cas9 constructs to knock out each homolog in the ein3-1 eil1-1 mutant background to create higher order mutants to reveal their ethylene-related functions, if any. Thus, my project is expected to expand our limited knowledge of ethylene-triggered translational regulation, further illuminate the role of auxin in response to ethylene, and address the nature of the fast ethylene response by shedding light on the remaining EIN3/EIL family members.