The Transcriptional Regulator LEUNIG_HOMOLOG Regulates Mucilage Release from the Arabidopsis Testa

Murray Walker,Antony Bacic,Filomena A Pettolino,Muhammad Tehseen,John F Golz,Monika S Doblin,Sarah M Wilson

doi:10.1104/pp.111.172692

Murray Walker, Antony Bacic + Show 5 more

Open Access

https://doi.org/10.1104/pp.111.172692

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Abstract

Exposure of the mature Arabidopsis (Arabidopsis thaliana) seed to water results in the rapid release of pectinaceous mucilage from the outer cells of the testa. Once released, mucilage completely envelops the seed in a gel-like capsule. The physical force required to rupture the outer cell wall of the testa comes from the swelling of the mucilage as it expands rapidly following hydration. In this study, we show that mutations in the transcriptional regulator LEUNIG_HOMOLOG (LUH) cause a mucilage extrusion defect due to altered mucilage swelling. Based on sugar linkage and immunomicroscopic analyses, we show that the structure of luh mucilage is altered, having both an increase in substituted rhamnogalacturonan I and in methyl-esterified homogalacturonan. Also correlated with the structural modification of luh mucilage is a significant decrease in MUCILAGE MODIFIED2 (MUM2; a β-galactosidase) expression in the luh seed coat, raising the possibility that reduced activity of this glycosidase is directly responsible for the luh mucilage defects. Consistent with this is the structural similarity between mum2 and luh mucilage as well as the observation that elevating MUM2 expression in luh mutants completely suppresses the mucilage extrusion defect. Suppression of the luh mutant phenotype was also observed when LEUNIG, a transcriptional corepressor closely related to LUH, was introduced in luh mutants under the control of the LUH promoter. Based on these data, we propose a new model for the regulation of pectin biosynthesis during plant growth and development.

Highlights

Exposure of the mature Arabidopsis (Arabidopsis thaliana) seed to water results in the rapid release of pectinaceous mucilage from the outer cells of the testa
Given that the enhanced extrusion defect of the luh; lug/+ seeds was not associated with an obvious reduction in mucilage accumulation within the mucilage-secreting cells (MSC) (Supplemental Fig. S5), we addressed whether this phenotype was correlated with a further decrease in MUCILAGE MODIFIED2 (MUM2) expression. quantitative reverse transcription (qRT)-PCR analysis, failed to detected a significant expression difference
This study has established that mutations in LUH, a gene closely related to the transcriptional corepressor LUG (Liu and Karmarkar, 2008), affect mucilage release from the seed coat following contact with water

Summary

RESULTS

Luh Mutants Display a Mucilage Extrusion Defect Associated with Altered Mucilage Hydration Properties. In addition to having similar mucilage extrusion defects, the MSC of luh and mum have slightly thicker radial cell walls (Fig. 3, A–D) To test whether these phenotypic similarities are a consequence of LUH and MUM2 functioning in the same genetic pathway, we used quantitative reverse transcription (qRT)-PCR to examine the expression of MUM2 in developing wild-type and luh mutant seeds. Subsequent qRT-PCR analysis confirmed that MUM2 expression was close to wild-type levels in 10-dpa seed tissue obtained from two independent class I luh;LUHpro::MUM2 lines (Table III). This confirms that elevating MUM2 expression in luh mutants restores RG-I substitution to wild-type levels and reestablishes mucilage extrusion. C and D, Ruthenium red-stained seeds from lug mutant (C) and luh;LUHpro::LUG transgenic (D) plants displaying wild-type levels of mucilage release. Consistent with redundancy between these coregulators, RT-PCR analysis detected expression of all four genes in these tissues (Fig. 7G)

DISCUSSION

Findings

MATERIALS AND METHODS