SPX1 is a phosphate-dependent inhibitor of PHOSPHATE STARVATION RESPONSE 1 in Arabidopsis

Abstract

To cope with growth in low-phosphate (Pi) soils, plants have evolved adaptive responses that involve both developmental and metabolic changes. PHOSPHATE STARVATION RESPONSE 1 (PHR1) and related transcription factors play a central role in the control of Pi starvation responses (PSRs). How Pi levels control PHR1 activity, and thus PSRs, remains to be elucidated. Here, we identify a direct Pi-dependent inhibitor of PHR1 in Arabidopsis, SPX1, a nuclear protein that shares the SPX domain with yeast Pi sensors and with several Pi starvation signaling proteins from plants. Double mutation of SPX1 and of a related gene, SPX2, resulted in molecular and physiological changes indicative of increased PHR1 activity in plants grown in Pi-sufficient conditions or after Pi refeeding of Pi-starved plants but had only a limited effect on PHR1 activity in Pi-starved plants. These data indicate that SPX1 and SPX2 have a cellular Pi-dependent inhibitory effect on PHR1. Coimmunoprecipitation assays showed that the SPX1/PHR1 interaction in planta is highly Pi-dependent. DNA-binding and pull-down assays with bacterially expressed, affinity-purified tagged SPX1 and ΔPHR1 proteins showed that SPX1 is a competitive inhibitor of PHR1 binding to its recognition sequence, and that its efficiency is highly dependent on the presence of Pi or phosphite, a nonmetabolizable Pi analog that can repress PSRs. The relative strength of the SPX1/PHR1 interaction is thus directly influenced by Pi, providing a link between Pi perception and signaling.phosphate sensor | phosphate starvation signaling S ince the beginning of molecular genetics, phosphate (Pi) starvation rescue systems, especially the Pi starvation rescues systems of bacteria and yeast, have served as emblematic models for studies of regulation of gene activity. In plants, these systems have gained additional interest because of the complexity and multicellular nature of plants (1, 2), and especially due to their potential for improving Pi acquisition and use in crops, a major goal toward sustainable agriculture. Considerable information has been gathered in the past decade on the components of the Pi starvation signaling pathway (reviewed in refs. 3-6). Major findings in plants include (i) identification of PHOSPHATE STARVATION RESPONSE 1 (PHR1) and related transcription factors as master regulators of Pi starvation responses (PSRs) (7-11); (ii) demonstration of the involvement of ubiquitin system components, including PHO2 and NLA, in Pi signaling (12-16); (iii) identification of miRNAs as mobile signals in Pi homeostasis (17, 18); and (iv) identification of Pi starvation-induced (PSI) riboregulators of miRNA activity, based on target mimicry (19) and natural antisense RNA that activates translation of PHO1 mRNA (20). In addition, a singular characteristic of nutrient starvation responses in plants is that several of these responses are at long distance, systemically controlled by plant shoot nutrient status, whereas others are controlled by local nutrient concentration. Transcriptomic a...

Methylome analysis reveals an important role for epigenetic changes in the regulation of the Arabidopsis response to phosphate starvation

“…Many significant advances have been achieved in our understanding of the signaling mechanisms that regulate the transcriptional activation of PSR genes, including the characterization of the central regulator PHR1 and its interaction with SPX1 and SPX2 repressors to control the activation of PSR genes (37). Our results showed that the mechanisms that regulate Arabidopsis responses to Pi starvation also involve changes in the DNA methylation status, thereby modulating the expression of PSR genes and root system developmental responses to low Pi availability.…”

“…Of particular interest is the finding that two key regulators of Arabidopsis responses to low Pi were among the Pf-methDEGs, namely SPX2, which inhibits PHOSPHATE STARVATION RESPONSE 1 (PHR1) DNA-binding activity in a Pi-dependent manner (37), and miR827 (38,39), which posttranscriptionally represses NITROGEN LIMITATION ADAPTATION (NLA) to release the negative regulation of PHOSPHATE TRANSPORTER1 (PHT1.1) and PHOSPHATE TRANSPORTER TRAFFIC FACILITATOR1 (PHF1), both of which are involved in Pi transport (39). The differential methylation changes observed in these two genes were highly consistent among tissues and over time under the LP conditions.…”

The Rice CK2 Kinase Regulates Trafficking of Phosphate Transporters in Response to Phosphate Levels

“…Thus, CK2, PHO2, and NLA generate a negative feedback loop for the regulation of PT activity according to plant Pi status. Such a negative loop is paralleled by a similar feedback mechanism for transcriptional control generated between the PHR1 master regulator of Pi starvation responses and several SPX proteins (Lv et al, 2014;Puga et al, 2014;Wang et al, 2014).…”

The Phosphate Fast-Responsive Genes PECP1 and PPsPase1 Affect Phosphocholine and Phosphoethanolamine Content

“…Furthermore, their corresponding mutants exhibit altered gene induction and classical Pi starvation responses, including diminished effects on Pi content and lipid remobilization (Pant et al, 2015). Candidates for the systemic control of Pi starvation responses have been proposed, including substantial transport of mRNAs throughout the plant (as reviewed in Puga et al, 2017), and the direct sensing of Pi concentration (or Pi-containing metabolites) by SPX-type proteins (Wang et al, 2009;Puga et al, 2014;Wild et al, 2016).Two phosphatases belonging to the haloacid dehalogenase (HAD) superfamily were identified as some of the most strongly induced transcripts following Pi starvation (Bari et al, 2006;Müller et al, 2007;Thibaud et al, 2010). These enzymes were named PPsPase1 (for pyrophosphatase [PPi]-specific phosphatase1) and PECP1 (for phosphoethanolamine (PEth)/ phosphocholine [PCho] phosphatase1) on the basis of their in vitro activity, following their heterologous expression in bacteria and subsequent purification (May et al, 2011(May et al, , 2012.…”

“…Furthermore, their corresponding mutants exhibit altered gene induction and classical Pi starvation responses, including diminished effects on Pi content and lipid remobilization (Pant et al, 2015). Candidates for the systemic control of Pi starvation responses have been proposed, including substantial transport of mRNAs throughout the plant (as reviewed in Puga et al, 2017), and the direct sensing of Pi concentration (or Pi-containing metabolites) by SPX-type proteins (Wang et al, 2009;Puga et al, 2014;Wild et al, 2016).…”