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Planta

, 229:593 | Cite as

BREVIS RADIX is involved in cytokinin-mediated inhibition of lateral root initiation in Arabidopsis

  • Jing Li
  • Xiaorong Mo
  • Jirong Wang
  • Nannan Chen
  • Huan Fan
  • Chunyan Dai
  • Ping Wu
Original Article

Abstract

In contrast to auxin, relatively little is known about the molecular mechanism of cytokinin (CTK) inhibition of lateral root initiation. Previous studies demonstrated that BREVIS RADIX (BRX), a protein of unknown biochemical function, maintains a rate-limiting brassinosteroid biosynthesis enzyme expression to keep brassinosteroid biosynthesis above a critical threshold. Here, we show that the brx-2 mutant is insensitive to exogenous CTK-induced inhibition of lateral root initiation and that this can be restored by embryonic brassinosteroid treatment. However post-embryonic brassinosteroid treatment can not rescue brx-2 mutant phenotype in the presence of CTK. Meanwhile the brassinosteroid receptor defective mutant bri1-6 shows normal CTK-mediated inhibition on LR growth. These results suggest the CTK-mediated inhibition of LR initiation is not directly dependent on brassinosteroid level. Furthermore, compared with wild type, brx-2 exhibits altered auxin response in presumptive founder cells, lateral root primodia and primary root tip in the presence of exogenous CTK. We concluded that CTK inhibition on lateral root initiation depend on specific auxin response loss in presumptive founder cell. The aberrant primary root growth caused by the embryonic brassinosteroid shortage can indirectly result in the lateral root phenotype of brx-2 in presence of CTK.

Keywords

Arabidopsis Auxin Brassinosteroid BRX Cytokinin Lateral root initiation 

Abbreviations

BA

6-Benzylaminopurine

BL

Brassinolide

BRX

BREVIS RADIX

CPD

CONSTITUTIVE PHOTOMORPHOGENESIS AND DWARF

CTK

Cytokinin

DAG

Days after germination

IAA

Indole-3-acetic acid

LR

Lateral root

LRI

Lateral root initiation

LRP

Lateral root primordium

NAA

α-Naphthaleneacetic acid

QRT-PCR

Quantitative real-time PCR

Notes

Acknowledgments

We thank Professor Malcolm Bennett’s group of Nottingham University for their kindly providing the DR5::GUS and IAA2::GUS maker lines; Peter Doerner of University of Edinburgh for the CYCB1;1::GUS line; Jason W. Reed (University of North Carolina, Chapel Hill, NC, USA) for the axr2-1 seeds. The seeds of bri1-6 and sur1 are supplied by ABRC (Arabidopsis Biological Resource Center, Ohio State University, Columbus, OH, USA). This work was supported by the National Natural Science Foundation in China (30600046). We are grateful to Professor Hanma Zhang (University of Leeds, UK) for generously providing the brx-2 mutant seeds and Professor Peter Hedden (Rothamsted Research, UK) for reviewing this paper.

Supplementary material

425_2008_854_MOESM1_ESM.jpeg (537 kb)
Supplement Table. 1 The sequence information and the chromosomal positions of markers inmapping of brx-2. (537 KB)
425_2008_854_MOESM2_ESM.jpeg (183 kb)
Supplement Fig. 1 The brx-2 mutant show sensitive to CTK-induced responses anthocyanin accumulation (a) and chlorophyll retention assay (b). To determine the effect of CTK onanthocyanin accumulation, seeds were plated on MS medium supplemented with 5 μM BA. After 10days of growth, the seedlings were weighed and anthocyanin was extracted and estimated. Anthocyanin was extracted from seedlings by overnight extraction in 3 ml of 1% (v/v) acidic methanol. Phase partitioning was done next day by adding 3 ml chloroform and 2 ml reverse-osmosiswater. Absorbance in the aqueous phase was recorded spectrophotometrically at 530nm. For chlorophyll retention assay, fully expanded leaves were excised from 24-day-old plants (initial) and floated on water supplemented with 0.1% DMSO carrier control or BA for 7 d in the dark.Chlorophyll was extracted and quantified spectrophotometrically from freshly cut leaves and senesced leaves as in the seedling chlorophyll analysis. Absorbance of aqueous phase was recordedspectrophotometrically at 652nm. (183 KB)
425_2008_854_MOESM3_ESM.jpeg (938 kb)
Supplement Fig. 2 RT-PCR analysis for transcription of the genes for CTK biosynthesis and signalling pathway. 20 DAG seedlings of the WT and brx-2 mutant grown on control medium and then transferred to control or 10 μM BA medium for 24 hours. (938 KB)
425_2008_854_MOESM4_ESM.jpeg (49 kb)
Supplement Fig. 3 The brx-2 mutant is sensitive to auxin. Comparison of the number of visible LRs between WT and brx-2 seedlings on 0.1 μM IAA and 0.1 μM NAA medium (n=30). (48.5 KB)

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Copyright information

© Springer-Verlag 2008

Authors and Affiliations

  • Jing Li
    • 1
  • Xiaorong Mo
    • 1
  • Jirong Wang
    • 1
  • Nannan Chen
    • 1
  • Huan Fan
    • 1
  • Chunyan Dai
    • 1
  • Ping Wu
    • 1
  1. 1.State Key Laboratory of Plant Physiology and Biochemistry, College of Life SciencesZhejiang UniversityHangzhouPeople’s Republic of China

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