First published online September 3, 2004; 10.1104/pp.104.041665
Plant Physiology 136:2747-2761 (2004)
© 2004 American Society of Plant Biologists
CELL BIOLOGY AND SIGNAL TRANSDUCTION
Two Transcription Factors Are Negative Regulators of Gibberellin Response in the HvSPY-Signaling Pathway in Barley Aleurone[w]
Masumi Robertson*
Commonwealth Scientific and Industrial Research Organisation Plant Industry, Canberra, Australian Capitol Territory 2601, Australia
SPINDLY (SPY) protein from barley (Hordeum vulgare L. cv Himalaya; HvSPY) negatively regulated GA responses in aleurone, and genetic analyses of Arabidopsis thaliana predict that SPY functions in a derepressible GA-signaling pathway. Many, if not all, GA-dependent responses require SPY protein, and to improve our understanding of how the SPY signaling pathway operates, a yeast two-hybrid screen was used to identify both upstream and downstream components that might regulate the activity of the HvSPY protein. A number of proteins from diverse classes were identified using HvSPY as bait and barley cDNA libraries as prey. Two of the HvSPY-interacting (HSI) proteins were transcription factors belonging to the myb and NAC gene families, HSImyb and HSINAC. Interaction occurred via the tetratricopeptide repeat domain of HvSPY and specificity was shown both in vivo and in vitro. Messenger RNAs for these proteins were expressed differentially in many parts of the barley plant but at very low levels. Both HSImyb and HSINAC inhibited the GA3 up-regulation of  -amylase expression in aleurone, both were activators of transcription in yeast, and the green fluorescent protein-HSI fusion proteins were localized in the nucleus. These results are consistent with the model that HSI transcription factors act downstream of HvSPY as negative regulators and that they in turn could activate other negative regulators, forming the HvSPY negative regulator-signaling pathway for GA response.
[w] The online version of this article contains Web-only data.
Article, publication date, and citation information can be found at www.plantphysiol.org/cgi/doi/10.1104/pp.104.041665.
* E-mail masumi.robertson{at}csiro.au; fax 61 (2) 6246–5000.
Received February 29, 2004;
returned for revision June 20, 2004;
accepted June 28, 2004.
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