The BRASSINOSTEROID INSENSITIVE1–LIKE3 Signalosome Complex Regulates Arabidopsis Root Development

Norma Fàbregas,Steven D Clouse,Michael B Goshe,Na Li,Ana I Caño-Delgado,Sjef Boeren,Tara E Nash,Sacco De Vries

doi:10.1105/tpc.113.114462

Abstract

Brassinosteroid (BR) hormones are primarily perceived at the cell surface by the leucine-rich repeat receptor-like kinase brassinosteroid insensitive1 (BRI1). In Arabidopsis thaliana, BRI1 has two close homologs, BRI1-LIKE1 (BRL1) and BRL3, respectively, which are expressed in the vascular tissues and regulate shoot vascular development. Here, we identify novel components of the BRL3 receptor complex in planta by immunoprecipitation and mass spectrometry analysis. Whereas BRI1 associated kinase1 (BAK1) and several other known BRI1 interactors coimmunoprecipitated with BRL3, no evidence was found of a direct interaction between BRI1 and BRL3. In addition, we confirmed that BAK1 interacts with the BRL1 receptor by coimmunoprecipitation and fluorescence microscopy analysis. Importantly, genetic analysis of brl1 brl3 bak1-3 triple mutants revealed that BAK1, BRL1, and BRL3 signaling modulate root growth and development by contributing to the cellular activities of provascular and quiescent center cells. This provides functional relevance to the observed protein-protein interactions of the BRL3 signalosome. Overall, our study demonstrates that cell-specific BR receptor complexes can be assembled to perform different cellular activities during plant root growth, while highlighting that immunoprecipitation of leucine-rich repeat receptor kinases in plants is a powerful approach for unveiling signaling mechanisms with cellular resolution in plant development.

Highlights

Plant steroid hormones brassinosteroids (BRs) are perceived by the plasma membrane–localized BRASSINOSTEROID INSENSITIVE1 (BRI1; Li and Chory, 1997)
Protein Expression of BRI1-Like Family Members In Arabidopsis, expression of BRL1 and BRL3 is enriched in the vascular tissues, whereas that of BRI1 appears in most plant cells (Caño-Delgado et al, 2004)
To reveal the localization of BRL1 and BRL3 receptors, the full-length genomic sequences of BRL1 and BRL3 were fused to yellow fluorescent protein (YFP) under the control of the native promoters consisting of a region 2 kb upstream of the start codon for BRL1 and BRL3 (ProBRL1: BRL1-YFP and ProBRL3:BRL3-YFP, respectively)

Summary

Introduction

Plant steroid hormones brassinosteroids (BRs) are perceived by the plasma membrane–localized BRASSINOSTEROID INSENSITIVE1 (BRI1; Li and Chory, 1997). Brassinolide (BL) binding occurs at the BRI1 extracellular domain. This domain consists of 25 LRRs interrupted by a 70–amino acid island domain between the 21st and 22nd LRR, which creates a surface pocket for ligand binding (Wang et al, 2001; Kinoshita et al, 2005; Hothorn et al, 2011; She et al, 2011). Recent evidence suggests that SERK coreceptors are essential for BRI1-mediated signaling (Gou et al., 2012). Downstream of BRI1 and SERKs, members of the BRASSINOSTEROID SIGNALING KINASE (BSK) cytoplasmic kinase family are subject to BRI1-mediated phosphorylation (Tang et al, 2008), and subsequently the signal is transmitted to BRI1-EMS SUPPRESSOR1 (BES1) and BRASSINAZOLE-RESISTANT1 (BZR1) transcription factors (Wang et al, 2002; Yin et al, 2005)

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Results

Conclusion