Arabidopsis WRKY45 Transcription Factor Activates PHOSPHATE TRANSPORTER1;1 Expression in Response to Phosphate Starvation

Hui Wang,Yi-Fang Chen,Jun-Ye Duan,Yun Chen,Wei-Hua Wu,Qian Xu,You-Han Kong

doi:10.1104/pp.113.235077

Hui Wang, Yi-Fang Chen + Show 5 more

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https://doi.org/10.1104/pp.113.235077

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Abstract

The WRKY transcription factor family has more than 70 members in the Arabidopsis (Arabidopsis thaliana) genome, and some of them are involved in plant responses to biotic and abiotic stresses. This study evaluated the role of WRKY45 in regulating phosphate (Pi) uptake in Arabidopsis. WRKY45 was localized in the nucleus and mainly expressed in roots. During Pi starvation, WRKY45 expression was markedly induced, typically in roots. WRKY45 overexpression in Arabidopsis increased Pi content and uptake, while RNA interference suppression of WRKY45 decreased Pi content and uptake. Furthermore, the WRKY45-overexpressing lines were more sensitive to arsenate, the analog of Pi, compared with wild-type seedlings. These results indicate that WRKY45 positively regulates Arabidopsis Pi uptake. Quantitative real-time polymerase chain reaction and β-glucuronidase staining assays showed that PHOSPHATE TRANSPORTER1;1 (PHT1;1) expression was enhanced in the WRKY45-overexpressing lines and slightly repressed in the WRKY45 RNA interference line. Chromatin immunoprecipitation and electrophoretic mobility shift assay results indicated that WRKY45 can bind to two W-boxes within the PHT1;1 promoter, confirming the role of WRKY45 in directly up-regulating PHT1;1 expression. The pht1;1 mutant showed decreased Pi content and uptake, and overexpression of PHT1;1 resulted in enhanced Pi content and uptake. Furthermore, the PHT1;1-overexpressing line was much more sensitive to arsenate than WRKY45-overexpressing and wild-type seedlings, indicating that PHT1;1 overexpression can enhance Arabidopsis Pi uptake. Moreover, the enhanced Pi uptake and the increased arsenate sensitivity of the WRKY45-overexpressing line was impaired by pht1;1 (35S:WRKY45-18::pht1;1), demonstrating an epistatic genetic regulation between WRKY45 and PHT1;1. Together, our results demonstrate that WRKY45 is involved in Arabidopsis response to Pi starvation by direct up-regulation of PHT1;1 expression.

Highlights

The WRKY transcription factor family has more than 70 members in the Arabidopsis (Arabidopsis thaliana) genome, and some of them are involved in plant responses to biotic and abiotic stresses
Quantitative real-time PCR analysis showed that WRKY45 was mainly expressed in the roots (Fig. 1A), and when wild-type Arabidopsis seedlings were challenged with Pi starvation, the WRKY45 was significantly induced, typically in the roots (Fig. 1A)
Seven-day-old ProWRKY45:GUS seedlings were transferred to Murashige and Skoog (MS) medium (Pi-sufficient condition, MS) or MS medium without Pi (Pi-deficient condition, low phosphate medium [LP]) for 5 d and stained to detect GUS activity

Summary

Introduction

The WRKY transcription factor family has more than 70 members in the Arabidopsis (Arabidopsis thaliana) genome, and some of them are involved in plant responses to biotic and abiotic stresses. The WRKY45-overexpressing lines were more sensitive to arsenate, the analog of Pi, compared with wild-type seedlings These results indicate that WRKY45 positively regulates Arabidopsis Pi uptake. Transcriptome analysis demonstrated that the expression of many genes, including PHT1 family genes, significantly changes in Oryza sativa (Wasaki et al, 2003) and Arabidopsis (Wu et al, 2003; Misson et al, 2005) in response to Pi starvation, indicating that transcriptional regulation plays important roles in plants’ responses to low-Pi stress. Several WRKY proteins, such as AtWRKY75 and AtWRKY6, have been reported to play important roles in plant responses to Pi starvation (Devaiah et al, 2007a; Chen et al, 2009)

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