Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2022 May 8;23(9):5241.
doi: 10.3390/ijms23095241.

One AP2/ERF Transcription Factor Positively Regulates Pi Uptake and Drought Tolerance in Poplar

Ningning Chen  1 , Jiajia Qin  1 , Shaofei Tong  1 , Weiwei Wang  1 , Yuanzhong Jiang  1
Affiliations

One AP2/ERF Transcription Factor Positively Regulates Pi Uptake and Drought Tolerance in Poplar

Ningning Chen et al. Int J Mol Sci. .

Abstract

Drought decreases the inorganic phosphate (Pi) supply of soil, resulting in Pi starvation of plants, but the molecular mechanism of how plants, especially the perennial trees, are tolerant to drought stress and Pi starvation, is still elusive. In this study, we identified an AP2/ERF transcription factor gene, PalERF2, from Populus alba var. pyramidalis, and it was induced by both mannitol treatment and Pi starvation. Overexpressing and knocking-down of PalERF2 both enhanced and attenuated tolerance to drought stress and Pi deficiency compared to WT, respectively. Moreover, the overexpression of PalERF2 up-regulated the expression levels of Pi starvation-induced (PSI) genes and increased Pi uptake under drought conditions; however, its RNAi poplar showed the opposite phenotypes. Subsequent analysis indicated that PalERF2 directly modulated expressions of drought-responsive genes PalRD20 and PalSAG113, as well as PSI genes PalPHL2 and PalPHT1;4, through binding to the DRE motifs on their promoters. These results clearly indicate that poplars can recruit PalERF2 to increase the tolerance to drought and also elevate Pi uptake under drought stress.

Keywords: PalERF2; Populus; drought stress; inorganic phosphate starvation; transcriptional regulation.

PubMed Disclaimer

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Characteristics of PalERF2 of Populus alba var. Pyramidalis. (A) Multiple sequence alignment of ERFs and the accession numbers are derived from different species (NM_001325036.1 from Nicotiana tabacum, XP_015636763.1 from rice, AT5G47220.1 from Arabidopsis thaliana, KAB5574006.1 from Salix brachista, and XP_003538752.2 from Glycine max). (B) The qRT-PCR analysis of PalERF2 expression in root (R), stem (S), mature leaves (ML), young leaves (YL), and petiole (P) in the mediums of MS. (C,D) The temporal expression pattern of PalERF2 under low Pi treatment (10 μM Pi) and 150 mM mannitol treatment in shoot and root, respectively. Error bars indicate SD values from three biological replicates. (E) Subcellular localization of PalERF2 in the mesophyll protoplasts of P. alba var. pyramidalis. The empty vector pBI221-expressing GFP is used as control (upper row) and the PalERF2-GFP fusion proteins are localized in the nucleus only (lower row). DAPI staining indicates the nucleus.
Figure 2
Figure 2
The PalERF2 transgenic poplars under low Pi condition. (A) Phenotypes of transgenic and WT poplars grew in liquid medium with 10 μM Pi for 4 weeks. (B) The Pi contents of transgenic and WT poplars in root and shoot after low Pi treatment. (C) Anthocyanin contents of WT and transgenic poplar after low Pi treatment. Error bars indicate SD values from five biological replicates. Significant differences were analyzed by Duncan’s test (p < 0.05, n = 5). Different letters indicate statistically significant differences. (D) The qRT-PCR analysis of Pi starvation response (PSR) genes in PalERF2-OE, PalERF2-RNAi, and WT poplars. Error bars indicate SD values from three biological replicates.
Figure 3
Figure 3
PalERF2 regulates PalPHL2 and PalPHT1;4 expression. (A) Structures of effector and reporters employed in dual-luciferase assay. (B) Transient co-expression of effector and reporter vectors in Nicotiana benthamiana leaves for dual-luciferase assay. Error bars indicate SD values (n = 3). Asterisks indicate significant differences compared to control by Student’s t-test, ***, p < 0.01. (C) Distribution of core DRE motifs in the promoters of PalPHL2 and PalPHT1;4. (D,E) ChIP-qPCR determined the binding of PalERF2 to the PalPHL2 and PalPHT1;4 promoter regions containing DRE, respectively. Error values represent means ± SD (n = 3). Significant differences were analyzed by Duncan’s test (p < 0.05, n = 5). Different letters indicate statistically significant differences. (F) EMSA tested the binding activity of PalERF2 to the DRE in PalPHL2 and PalPHT1;4 promoters. The unlabeled cold probes were added to compete with labeled probes. + means the cold probe is 20 times the labeled probe, ++ means 50 times. The arrows mark the binding probe and free probe.
Figure 4
Figure 4
The phenotypes of PalERF2 transgenic poplars under drought stress. (A) The phenotypes of transgenic and WT poplars after 5 days of drought treatment. (B) The MDA contents were measured after drought treatment. (C) The total chlorophyll contents were measured after drought treatment. (B,C) values represents means ± SD (n = 5). Significance of differences was analyzed by Duncan’s test (p < 0.05, n = 5). Different letters indicate statistically significant difference. (D) The relative expression of drought-associated genes in PalERF2-OE, PalERF2-RNAi, and WT poplars. Error bars indicate SD values from three biological replicates.
Figure 5
Figure 5
PalERF2 directly regulated the expression of PalRD20 and PalSAG113. (A) Structures of effector and reporters employed in Dual-luciferase assay. (B) Transient co-expression of effector and reporter vectors in N. benthamiana leaves. Data shown as mean ± SD (n = 3). Asterisks indicate significant differences compared to control by Student’s t-test, ***, p < 0.01. (C) Distribution of DRE and core DRE motifs in the promoter of PalRD20 and PalSAG113. (D,E) ChIP-qPCR demonstrated that PalERF2 bound to the promoter region of PalRD20 and PalSAG113 containing DRE in vivo. Significant differences were analyzed by Duncan’s test (p < 0.05, n = 5). Different letters indicate statistically significant differences. (F) EMSA demonstrated that PalERF2 bound to the DRE in the PalRD20 and PalSAG113 promoters. Unlabeled cold probes as a competitor to compete with labeled probes. + means the cold probe is 20 times the labeled probe, ++ means 50 times. The arrows mark the binding probe and free probe.
Figure 6
Figure 6
The Pi contents and the expression of PSR genes in PalERF2 transgenic and WT poplars. (A) The Pi contents in WT and transgenic plants before drought treatment. (B) The Pi contents in WT and transgenic poplars after drought treatment. (A,B) Error bars indicate SD values from three biological replicates. Significant difference was analyzed by Duncan’s test (p < 0.05, n = 5). Different letters indicate statistically significant differences. (C) The qRT-PCR analyzed the relative expression of PSR genes in WT and transgenic poplars after drought treatment. Error bars indicate SD values from three biological replicates.
Figure 7
Figure 7
The proposed model for the PalERF2 mediated drought stress and low Pi responses in poplars.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. López-Bucio J., Hernández-Abreu E., Sánchez-Calderón L., Nieto-Jacobo M.F., Simpson J., Herrera-Estrella L. Phosphate availability alters architecture and causes changes in hormone sensitivity in the Arabidopsis root system. Plant Physiol. 2002;129:244–256. doi: 10.1104/pp.010934. - DOI - PMC - PubMed
    1. Franco-Zorrilla J.M., Martin A.C., Leyva A., Par-Ares J.P. Interaction between phosphate-starvation, sugar and cytokinin signaling in and the roles of cytokinin receptors CRE1/AHK4 and AHK3. Plant Physiol. 2005;138:847–857. doi: 10.1104/pp.105.060517. - DOI - PMC - PubMed
    1. Jain A., Nagarajan V.K., Raghothama K.G. Transcriptional regulation of phosphate acquisition by higher plants. Cell Mol. Life Sci. 2012;69:3207–3224. doi: 10.1007/s00018-012-1090-6. - DOI - PMC - PubMed
    1. Rennenberg H., Herschbach C. Phosphorus nutrition of woody plants: Many questions-few answers. Plant Biol. 2013;15:785–788. doi: 10.1111/plb.12078. - DOI - PubMed
    1. Staaf H. Plant nutrient changes in beech leaves during senescence as influenced by site characteristics. Acta. Oecol. Oec. Plant. 1982;3:161–170.

MeSH terms