A role for two-component signaling elements in the Arabidopsis growth recovery response to ethylene

Abstract

Previous studies indicate that the ability of Arabidopsis seedlings to recover normal growth following an ethylene treatment involves histidine kinase activity of the ethylene receptors. As histidine kinases can function as inputs for a two-component signaling system, we examined loss-of-function mutants involving two-component signaling elements. We find that mutants of phosphotransfer proteins and type-B response regulators exhibit a defect in their ethylene growth recovery response similar to that found with the loss-of-function ethylene receptor mutant etr1-7. The ability of two-component signaling elements to regulate the growth recovery response to ethylene functions independently from their well-characterized role in cytokinin signaling, based on the analysis of cytokinin receptor mutants as well as following chemical inhibition of cytokinin biosynthesis. Histidine kinase activity of the receptor ETR1 also facilitates growth recovery in the ethylene hypersensitive response, which is characterized by a transient decrease in growth rate when seedlings are treated continuously with a low dose of ethylene; however, this response was found to operate independently of the type-B response regulators. These results indicate that histidine kinase activity of the ethylene receptor ETR1 performs two independent functions: (a) regulating the growth recovery to ethylene through a two-component signaling system involving phosphotransfer proteins and type-B response regulators and (b) regulating the hypersensitive response to ethylene in a type-B response regulator independent manner.

Keywords: Arabidopsis; ethylene; histidine kinase; phosphotransfer protein; receptor; response regulator.

Figure 1

Growth kinetics of two‐day‐old etiolated Arabidopsis hypocotyls containing mutations in two‐component signaling elements. Short‐term growth kinetic analysis in response to 10 μl/L ethylene was performed on mutants involving receptors (column 1), AHPs (column 2), and type‐B ARRs (column 3) and compared to the wild type. Ethylene was introduced one hour after measurements were initiated (down arrow) and then removed two hours later (up arrow). For receptor mutants, we examined the etr1‐7 mutant as well as ahk double mutants. For ahp mutants, we examined ahp5‐2, ahp2 ahp3, and ahp2 ahp3 ahp5‐2. For type‐B arr mutants, we examined arr2‐4, arr10‐2 arr12‐1, and arr2‐2 arr10‐2 arr12‐1. Error bars indicate SE (n ≥ 3)

Figure 2

Hypocotyl growth response of two‐component mutants to cytokinin. Dose–response for the effect of the cytokinin t‐zeatin on hypocotyl elongation of four‐day‐old etiolated seedlings, performed as described (Argyros et al., 2008). The hypocotyl length of each line in the presence of cytokinin is expressed as a percentage of the control. Error bars indicate SE (n ≥ 9); error bars not shown if smaller than symbol

Figure 3

Lovastatin does not affect growth kinetics of two‐day‐old etiolated Arabidopsis hypocotyls. Short‐term growth kinetic analysis in response to 10 μl/L ethylene was performed on wild‐type seedlings grown in the absence (DMSO control) or the presence of 1 μM lovastatin. Data were normalized to growth rate in air prior to treatment with ethylene to facilitate the comparison. Unnormalized growth rates in air were 0.33 ± 0.01 mm/hr for the control and 0.24 ± 0.01 mm/hr for the lovastatin‐treated seedlings. Error bars indicate SE (n ≥ 3)

Figure 4

Kinetic analysis of the hypersensitive response to ethylene. Growth kinetic analysis was performed with 8.7 nl/L ethylene introduced 1 hr after measurements were initiated (down arrow). The hypersensitive response of etr1‐7 (a), arr2‐2 arr10‐2 arr12‐1 (b), and transgenic etr1‐7 ers1‐2 lines containing wild‐type ETR1 (gETR1), or kinase‐inactive ETR1 (getr1‐HGG) (c) are all compared to wild type. The dotted line in panel C shows data for wild type for comparison. Data were normalized to growth rate in air prior to treatment with ethylene to facilitate comparisons. Error bars indicate SE (n ≥ 4)

Figure 5

Model for ethylene signal transduction. Signal output from the ethylene receptors diverges to regulate the CTR1/EIN2/EIN3 and AHP/ARR pathways. ETR1 is shown here as a representative ethylene receptor that has histidine kinase activity. Ethylene responses may be dependent on an individual pathway or have varying contributions from both pathways