Cloning, Characterization and Expression Analysis of the Phosphate Starvation Response Gene, <i>ClPHR1,</i> from Chinese Fir

• Main Authors: Wanting Chen, Ranhong Chen, Ying Zhang, Jiaoyang Li, Mulualem Tigabu, Xiangqing Ma, Ming Li
• Format: Article
• Language: English
• Published: MDPI AG 2020-01-01
• Series: Forests
• Subjects: <i>cunninghamia lanceolata</i>; <i>phr1</i>; pi utilization efficiency; quantitative real-time pcr
• Online Access: https://www.mdpi.com/1999-4907/11/1/104

The study on the function and sequence of <i>PHR1</i> (Phosphate Starvation Response gene 1) gene, which plays a central role in plant phosphorus (Pi) signal regulatory network, is of great significance to further study response mechanisms to Pi deficiency. In this work, the previously selected Pi-efficient Chinese fir clone M32 was used as research material to obtain the full-length sequence of <i>ClPHR1</i> transcription factors in Chinese fir by RACE (Rapid Amplification of cDNA Ends) full-length cloning technique, and the structure, function and subcellular localization of <i>ClPHR1</i> gene encoding protein were analyzed. The temporal and spatial expression characteristics of <i>ClPHR1</i> transcription factors in Chinese fir under low Pi stress were also analyzed, and the overexpression of <i>ClPHR1</i> gene in transgenic <i>Arabidopsis thaliana</i> was obtained to verify the function of <i>ClPHR1</i> gene under low Pi stress. The results showed that the length of the <i>ClPHR1</i> gene obtained by rapid amplification of cDNA ends technique was 1954 bp, of which 1512 bp was an open reading frame. <i>ClPHR1</i> was predicted to be an unstable hydrophilic protein with only one possible transmembrane domain. The <i>ClPHR1</i> gene had a highly conserved MYB-CC domain, which is similar to the <i>PHR1</i> gene of other plants. Phylogenetic tree analysis showed that the sequence had high homology with <i>PHR1</i> genes in the <i>Prunus</i> species. The <i>ClPHR1</i> was expressed in all organs of Chinese fir, with the highest expression in the roots, followed by the leaves with the lowest expression in stems. <i>ClPHR1</i> expression in roots was reduced dramatically at the beginning of Pi stress treatment and followed by an increase at 7days; in leaves, it increased dramatically at the beginning of Pi starvation treatment and showed a decreasing trend after 3 days; in stems, the expression level of <i>ClPHR1</i> increased after 7 days of Pi stress treatment. The transient expression vector was introduced into plant cells, and it was found that <i>ClPHR1</i> was located in the nucleus and was a MYB-CC transcription factor expressed in the cell nucleus. The <i>ClPHR1</i> overexpression vector was constructed, and then introduced into <i>Arabidopsis thaliana</i> by agrobacterium infection inflorescence method. The expressions of Pi transporter genes, <i>AtPHT1;1, AtPHT1;2, AtPHT1;8</i> and <i>AtPHT1;9</i>, was significantly higher in the overexpressing strain than that in the wild type strain. The results suggest that the <i>ClPHR1</i> transcription factor could regulate the regulation of downstream Pi transporter gene and increase Pi utilization efficiency of the Chinese fir under Pi stress.