Planta

, Volume 229, Issue 3, pp 523–537 | Cite as

Functional analysis of HvSPY, a negative regulator of GA response, in barley aleurone cells and Arabidopsis

  • Fiona Filardo
  • Masumi Robertson
  • Davinder Pal Singh
  • Roger W. Parish
  • Stephen M. Swain
Original Article
First Online:
Received:
Accepted:

Abstract

SPINDLY (SPY) is an important regulator of plant development, and consists of an N-half tetratricopeptide repeat (TPR) domain containing 10 TPR motifs and a C-half catalytic domain, similar to O-GlcNAc transferase (OGT) of animals. The best characterised role of SPY is a negative regulator of GA signalling, and all known spy alleles have been isolated based on increased GA response. Of the eight alleles that directly affect the TPR domain, all alter TPRs 6, 8 and/or 9. To test the hypothesis that a subset of TPRs, including 6, 8 and 9, are both essential and sufficient for the regulation of GA response, we overexpressed the full-length barley (Hordeum vulgare L.) SPY protein (HvSPY) and several deletion mutants in barley aleurone cells and in Arabidopsis wild type (WT) and spy-4 plants. Transient assays in barley aleurone cells, that also express endogenous HvSPY, demonstrated that introduced HvSPY and HvTPR inhibited GA3-induced α-amylase expression. With the exception of HvSPYΔ1–5, the other deletion proteins were partially active in the barley assay, including HvSPYΔ6–9 which lacks TPRs 6, 8 and 9. In Arabidopsis, analysis of seed germination under a range of conditions revealed that 35S:HvSPY increased seed dormancy. Hvspy-2, which lacks parts of the eighth and ninth TPRs, was able to partially complement all aspects of the spy-4 phenotype. In the presence of AtSPY, 35S:HvTPR caused some phenotypes consistent with a decrease in GA signalling, including increased seed sensitivity to paclobutrazol and delayed flowering. These plants also possessed distorted leaf morphology and altered epidermal cell shape. Thus, despite genetic analysis demonstrating that TPRs 6, 8 and 9 are required for regulation of GA signalling, our results suggest that these TPRs are neither absolutely essential nor sufficient for SPY activity.

Keywords

Arabidopsis Gibberellin Hordeum Tetratricopeptide repeat (TPR) domain SPINDLY Seed dormancy Plant development 

Abbreviations

GA

Gibberellin

GA3

Gibberellic acid

O-GlcNAc

N-acetylglucosamine

OGT

O-GlcNAc transferase

PAC

Paclobutrazol

SPY

SPINDLY

TPR

Tetratricopeptide repeat

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Notes

Acknowledgments

We thank Angelica Jermakow, Carol Sigston, Warren Hudson-Taylor and Judy Radik (CSIRO, Australia) for their technical support and Neil Olszewski (University of Minnesota, USA) for the Arabidopsis spy seeds. This work was supported by CSIRO and La Trobe University as part of a Riverlink PhD studentship.

Supplementary material

425_2008_843_MOESM1_ESM.doc (44 kb)
Supplemental Table 1 (DOC 43 kb)

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Copyright information

© Government Employee 2008

Authors and Affiliations

  • Fiona Filardo
    • 1
    • 3
  • Masumi Robertson
    • 2
  • Davinder Pal Singh
    • 1
  • Roger W. Parish
    • 3
  • Stephen M. Swain
    • 1
  1. 1.CSIRO Plant Industry, P.M.BMerbeinAustralia
  2. 2.CSIRO Plant IndustryCanberraAustralia
  3. 3.Department of BotanyLa Trobe UniversityBundooraAustralia

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