Abstract
Inorganic orthophosphate (Pi) is an essential nutrient for plant growth, and its availability strongly impacts crop yield. PHOSPHATE1 (PHO1) transfers Pi from root to shoot via Pi export into root xylem vessels. In this work, we demonstrate that an upstream open reading frame (uORF) present in the 5' untranslated region of the Arabidopsis (Arabidopsis thaliana) PHO1 inhibits its translation and influences Pi homeostasis. The presence of the uORF strongly inhibited the translation of a PHO1 5'UTR-luciferase construct in protoplasts. A point mutation removing the PHO1 uORF (ΔuORF) in transgenic Arabidopsis resulted in increased association of its mRNA with polysomes and led to higher PHO1 protein levels, independent of Pi availability. Interestingly, deletion of the uORF led to higher shoot Pi content and was associated with improved shoot growth under low external Pi supply and no deleterious effects under Pi-sufficient conditions. We further show that natural accessions lacking the PHO1 uORF exhibit higher PHO1 protein levels and shoot Pi content. Increased shoot Pi content was linked to the absence of the PHO1 uORF in a population of F2 segregants. We identified the PHO1 uORF in genomes of crops such as rice (Oryza sativa), maize (Zea mays), barley (Hordeum vulgare), and wheat (Triticum aesativum), and we verified the inhibitory effect of the rice PHO1 uORF on translation in protoplasts. Our work suggests that regulation of PHO1 expression via its uORF might be a genetic resource useful-both in natural populations and in the context of genome editing-toward improving plant growth under Pi-deficient conditions.
Highlights
PHOSPHATE1 (PHO1) transfers Pi from root to shoot via Pi export into root xylem vessels
PHO1 is only weakly regulated at the transcriptional level by Pi deficiency, while its protein stability is controlled by ubiquitination via PHO2 (Liu et al, 2012). pho2 mutants show enhanced expression of both PHOSPHATE TRANSPORTER1 (PHT1) and PHO1 proteins, and exhibit a constitutively active phosphate-deficiency starvation response, leading to reduced growth associated with excessive shoot Pi content (Aung et al, 2006; Bari et al, 2006)
We demonstrated that the Arabidopsis PHO1 mRNA harbors a small upstream open reading frame in the 59 untranslated region (UTR) that inhibits its translation
Summary
PHOSPHATE1 (PHO1) transfers Pi from root to shoot via Pi export into root xylem vessels. Pho mutants show enhanced expression of both PHT1 and PHO1 proteins, and exhibit a constitutively active phosphate-deficiency starvation response, leading to reduced growth associated with excessive shoot Pi content (Aung et al, 2006; Bari et al, 2006). Root expression of the PHO1 EXS domain alone, which lacks Pi transport activity, was enough to rescue the stunted shoot growth of pho mutants, despite shoot Pi levels remaining low (Wege et al, 2016). Both these studies showed that low Pi content can be dissociated from its main effect on growth and that PHO1 could be involved in modulating the Pi-deficiency signaling cascade and its effect on growth. While ectopic overexpression of PHO1 in leaves has been shown to lead to stunted growth associated with aberrant Pi export in leaf apoplasts (Stefanovic et al, 2011), the effects of enhanced PHO1 expression in its endogenous root tissues have not been studied with respect to Pi homeostasis and growth
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