Poster: Signaling, cell-to-cell

Abs # 448: Development of a model for ethylene receptor signaling based on in vitro mutagenesis and domain swapping between receptor isoforms

Presenter:	Wang, Wuyi , wuyiwang@wisc.edu
Authors	Wang, Wuyi  (A)   Esch, Jeff J (B)   Agula, Hasi  (C)   Bleecker, Anthony B (A)
Affiliations:	(A): Department of Botany and Laboratory of Genetics, University of Wisconsin-Madison (B): Department of Plant Biology, University of Minnesota (C): College of Life Sciences, Inner Mongolia University, P.R. China

In Arabidopsis, ethylene signaling is mediated by a family of five receptors. Biochemical and genetic studies of the ethylene receptor ETR1 have led to the development of a molecular model for signal/response coupling by the receptor. The ethylene-binding domain of ETR1 is modeled as three membrane-spanning helices which coordinate the ethylene-binding copper cofactor. Amino acid residues within the binding domain, which are conserved across plant and cyanobacterial lineages, occur along specific faces of the helices. Mutations in 37 residues were generated and mutant receptors transferred into yeast and receptor deficient Arabidopsis lines to assess the effects of the mutations on ethylene-binding activity and receptor output capacity, respectively. Mutations in 7 residues located primarily in the midregions of helix I and II eliminated ethylene binding and conferred dominant ethylene insensitivity in planta, potentially defining the ethylene-binding pocket of the receptor. Clustered mostly near the cytoplasmic ends of the helices I and III , a second major class of 13 mutants also conferred dominant ethylene insensitivity, but showed normal binding activity when expressed in yeast. A two-step model for signaling is proposed in which ethylene first binds to the copper cofactor, resulting in a subsequent shift of the receptor from a low-energy transmitter-on state to a high-energy transmitter-off state. Previous work indicated a special role for subfamily I receptors (ETR1 and ERS1) in signaling. Chimeric ETR1 receptors containing either an ethylene-binding domain, a GAF domain, or a histidine kinase-related domain from subfamily II failed to rescue an etr1;ers1 double mutant line, indicating that all three domains from subfamily I are required for subfamily I function.