结论:磷酸盐缺乏通过PHR1直接结合F3\'H和LDOX启动子上的P1BS基序促进拟南芥花色苷积累,从而上调其表达。磷是植物必需的元素之一,植物主要从土壤中吸收无机磷酸盐(Pi)。但是Pi缺乏是限制植物生长发育的常见因素。花青素在绿色组织(如叶片)中的积累是许多植物响应Pi饥饿的特点之一。然而,关于Pi饥饿诱导花青素积累的机制知之甚少。这里,我们发现磷酸盐饥饿反应基因1(PHR1)的突变,它编码参与拟南芥Pi饥饿信号传导的关键因素,在Pi限制条件下显着减弱花色苷的积累。此外,参与花青素生物合成的几个Pi缺乏上调基因的表达,如黄烷酮3'-羟化酶(F3'H),二氢黄酮醇4-还原酶(DFR),白花青素双加氧酶(LDOX),和生产花色苷色素1(PAP1),在phr1-1突变体中显著低于野生型(WT)。酵母单杂交(Y1H)分析和染色质免疫沉淀定量PCR(ChIP-qPCR)均表明PHR1可以与F3\'H和LDOX的启动子相互作用,而不是DFR和PAP1。通过电泳迁移率变化分析(EMSA),进一步证实,位于F3'H和LDOX启动子上的PHR1结合序列(P1BS)基序是PHR1结合所必需的。此外,在拟南芥原生质体中,PHR1增强了F3\'H和LDOX启动子的转录活性,但是当P1BS基序突变时,这些作用明显受损。一起来看,这些结果表明,PHR1正调节拟南芥中Pi饥饿诱导的花色苷积累,至少在某种程度上,通过直接结合位于启动子上的P1BS基序来上调花青素生物合成基因F3\'H和LDOX的转录。
CONCLUSIONS: Phosphate deficiency promotes anthocyanin accumulation in Arabidopsis through direct binding of PHR1 to the P1BS motifs on the promoters of F3\'H and LDOX and thereby upregulating their expression. Phosphorus is one of the essential elements for plants, and plants mainly absorb inorganic phosphate (Pi) from soil. But Pi deficiency is a common factor limiting plant growth and development. Anthocyanin accumulation in green tissues (such as leaves) is one of the characteristics of many plants in response to Pi starvation. However, little is known about the mechanism by which Pi starvation induces anthocyanin accumulation. Here, we found that the mutation of the gene PHOSPHATE STARVATION RESPONSE1 (PHR1), which encodes a key factor involved in Pi starvation signaling in Arabidopsis, significantly attenuates anthocyanin accumulation under Pi-limiting conditions. Moreover, the expression of several Pi deficiency-upregulated genes that are involved in anthocyanin biosyntheses, such as flavanone 3\'-hydroxylase (F3\'H), dihydroflavonol 4-reductase (DFR), leucoanthocyanidin dioxygenase (LDOX), and production of anthocyanin pigment 1 (PAP1), was significantly lower in the phr1-1 mutant than in the wild type (WT). Both yeast one-hybrid (Y1H) analysis and chromatin immunoprecipitation quantitative PCR (ChIP-qPCR) showed that PHR1 can interact with the promoters of F3\'H and LDOX, but not DFR and PAP1. By electrophoretic mobility shift assay (EMSA), it was further confirmed that the PHR1-binding sequence (P1BS) motifs located on the F3\'H and LDOX promoters are required for the PHR1 bindings. Also, in Arabidopsis protoplasts, PHR1 enhanced the transcriptional activity of the F3\'H and LDOX promoters, but these effects were markedly impaired when the P1BS motifs were mutated. Taken together, these results indicate that PHR1 positively regulates Pi starvation-induced anthocyanin accumulation in Arabidopsis, at least in part, by directly binding the P1BS motifs located on the promoters to upregulate the transcription of anthocyanin biosynthetic genes F3\'H and LDOX.