Structure-Function of the Ethylene Binding Domain
One topic of interest is to further define the structure and function of the ethylene binding domain. We have used both mutational and chemical analyses to uncover more information about how ethylene binds to the receptor and transduces the signal through the protein. Interestingly, ethylene receptors are metalloproteins containing copper ions. While copper is the natural co-factor, we have shown that the other group 11 transition metals (silver and gold) support ethylene binding to the receptors, whereas, other metals do not. This observation is of interest because silver, but not gold, blocks ethylene action in the plant. These observations led to the hypothesis that silver may block conformational changes in the receptor because it is larger than copper. Interestingly, silver mainly affects the ETR1 ethylene receptor, but not the other isoforms. Another chemical approach we have used is to analyze ethylene binding and receptor function in the presence of various strained alkenes and other small chemicals in order to better define the ethylene binding pocket, identify how copper is delivered to the receptors, and uncover useful chemicals that can be used agriculturally. Finally, we have used mutagenesis of conserved residues in the binding domain to define regions necessary for ethylene binding, turning the receptor off when ethylene binds, and maintaining a functional receptor. For more information refer to: Rodriguez et al., 1999; Wang et al., 2006; Binder et al., 2007; Pirrung et al., 2008; Binder et al., 2010; McDaniel and Binder, 2012; Li et al. 2017).

Growth responses in Arabidopsis
(click on image to link to time-lapse movies)

Seed germination during salt stress

Sub-functionalization of Ethylene Receptor Isoforms
A third area of current research is to determine the basis for ethylene receptor sub-functionalization (reviewed in Shakeel et al., 2013) . We have uncovered several instances where the receptors have become sub-functionalized. For instance, ETR1, ETR2, and EIN4 are required for normal growth recovery after removal of ethylene, while ERS1 and ERS2 are not. This function requires ETR1 histidine kinase activity (Binder et al., 2004). ETR1 plays the predominant role in mediating the inhibitory effects of silver ions on ethylene responses (McDaniel and Binder, 2012), in mediating many responses of roots to ethylene (Harkey et al., 2018) and in susceptibility to the cyst nematode Heterodera schactii (Piya et al., 2019). Interestingly, our research has also revealed instances of contrasting roles for certain receptor isoforms. The first instance that we found was that ETR1 is necessary and sufficient for ethylene-stimulated nutational bending (also called circumnutations) of Arabidopsis hypocotyls; by contrast loss of the other four receptor isoforms leads to constitutive nutations (Binder et al., 2006; Kim et al., 2011). Another instance of contrasting roles is that ETR1 (and to a lesser extent EIN4) inhibits and ETR2 stimulates seed germination during salt stress (Wilson et al., 2014a) and in darkness (Wilson et al., 2014b). The receiver domain of ETR1 has an important role during germination under salt stress. We've recently defined regions of the receiver domain important for specific traits (Bakshi et al., 2015). Thus, we have two instances where receptor isoforms have opposite roles in a trait. This is not explained by current models of ethylene signaling, but appears to involve crosstalk (see below).  We are now studying the mechanistic basis for these various roles.

Distribution of Putative Ethylene Receptors in Non-plant Species
(cyan- cyanobaceria, red- proteobacteria, yellow- other bacteria, purple- non-plant eukaryotes)