Association of the Arabidopsis CTR1 Raf-like kinase with the ETR1 and ERS ethylene receptors

Abstract

In Arabidopsis thaliana, signal transduction of the hormone ethylene involves at least two receptors, ETR1 and ERS, both of which are members of the two-component histidine protein kinase family that is prevalent in prokaryotes. The pathway also contains a negative regulator of ethylene responses, CTR1, which closely resembles members of the Raf protein kinase family. CTR1 is thought to act at or downstream of ETR1 and ERS based on double mutant analysis; however, the signaling mechanisms leading from ethylene perception to the regulation of CTR1 are unknown. By using the yeast two-hybrid assay, we detected a specific interaction between the CTR1 amino-terminal domain and the predicted histidine kinase domain of ETR1 and ERS. We subsequently verified these interactions by using an in vitro protein association assay(s). In addition, we determined that the amino-terminal domain of CTR1 can associate with the predicted receiver domain of ETR1 in vitro. Based on deletion analysis, the portion of CTR1 that interacts with ETR1 roughly aligns with the regulatory region of Raf kinases. These physical associations support the genetic evidence that CTR1 acts in the pathway of ETR1 and ERS and suggest that these interactions could be involved in the regulation of CTR1 activity.Ethylene has numerous effects on plant growth and development, such as fruit-ripening, organ abscission, seed germination, senescence, and the induction of certain defense responses (1). The cloning of genes corresponding to several ethylene-response mutants in Arabidopsis has begun to provide us with insight into the molecular basis of ethylene signal transduction (2-6). In Arabidopsis, there are at least two ethylene receptors, ETR1 and ERS, that are similar to each other. Plants appear to have multiple ethylene receptors; several different ETR1 and ERS homologs have been cloned recently from Arabidopsis (7) and tomato (8, 9). The ETR1 and ERS gene products are predicted to function as histidine protein kinases based on their sequence similarities with the two-component regulator family. The two-component regulators are prevalent in prokaryotes (10) and are starting to be identified in various eukaryotes, including fungi, slime mold, and higher plants (11,12); one of these, the Arabidopsis CKI1 protein, is potentially a receptor for cytokinin (13). The basic two-component system consists of a histidine autokinase sensor component that directs the activity of a cognate response regulator, which in turn controls downstream signaling (10). After autophosphorylation of the sensor kinase, the phosphoryl group is transferred from the histidine of the sensor kinase to an aspartic acid residue in the receiver domain of the response regulator (10). The ETR1 protein is an example of a hybrid histidine kinase because it contains both a histidine kinase domain and a receiver domain (2). ERS, in contrast, contains only a histidine kinase domain (3). The significance of having an attached receiver domain is unknown, and whether there are separate cognate...

Differential Control of Ethylene Responses byGREEN-RIPEandGREEN-RIPE LIKE1Provides Evidence for Distinct Ethylene Signaling Modules in Tomato

“…The ethylene receptors are known to form large heteromeric complexes, and interaction experiments between the ethylene receptors and additional signaling components suggest that these complexes likely contain AtRTE1, EIN2, and CTR1 and occur within the ER and Golgi membranes (Clark et al, 1998;Chen et al, 2002Gao et al, 2003Gao et al, , 2008Bisson et al, 2009;Dong et al, 2010). Our data indicate that SlGR, SlGRL1, and SlGRL2 are localized primarily to the Golgi membranes at steady state, although at present we cannot completely exclude the possibility that these proteins are also localized to additional organelles ( Fig.…”

“…Loss-of-function analyses indicate that the receptors act in a semiredundant manner to negatively regulate ethylene responses, although the ETR1 receptor appears to play more of a significant role in mediating ethylene responses than the other Arabidopsis receptors Cancel and Larsen, 2002;Hall and Bleecker, 2003;Wang et al, 2003;Binder et al, 2006;Qu et al, 2007). In the absence of ethylene, the receptors actively suppress downstream responses through direct binding of CONSTITUTIVE TRIPLE RESPONSE1 (CTR1), a Ser/Thr mitogen-activated protein kinase kinase kinase that acts as a negative regulator of the pathway (Kieber et al, 1993;Clark et al, 1998;Gao et al, 2003;Huang et al, 2003). Upon ethylene binding, a conformational change is thought to occur, leading to receptor inactivation and the release of CTR1-mediated suppression (Huang et al, 2003;Zhao and Guo, 2011).…”

Delayed Abscission and Shorter Internodes Correlate with a Reduction in the Ethylene Receptor LeETR1 Transcript in Transgenic Tomato

“…Also of interest in this regard is that, if the receiver domain was deleted from the AtETR1 protein, binding affinity between the AtETR1 and AtCTR1 proteins was much weakened (Clark et al, 1998). Moreover, the ability of the receiver domain to independently form dimers raises the possibility of interaction with other independent response regulators.…”

Modulation of Ethylene Responses Affects Plant Salt-Stress Responses

“…The ETR1 receptor function can be regulated by RTE1 or the copper transporter RAN1 Resnick et al, 2006). ETR1 and ERS1 can interact with CTR1 that may be modulated by protein phosphatase 2A (Clark et al, 1998;Gao et al, 2003;Larsen and Cancel, 2003). Homologous genes of ethylene receptors have been isolated from many other plants, e.g.…”