Comprehensive Genomic Identification and Expression Analysis of the Phosphate Transporter (PHT) Gene Family in Apple

Abstract

Elemental phosphorus (Pi) is essential to plant growth and development. The family of phosphate transporters (PHTs) mediates the uptake and translocation of Pi inside the plants. Members include five sub-cellular phosphate transporters that play different roles in Pi uptake and transport. We searched the Genome Database for Rosaceae and identified five clusters of phosphate transporters in apple (Malus domestica), including 37 putative genes. The MdPHT1 family contains 14 genes while MdPHT2 has two, MdPHT3 has seven, MdPHT4 has 11, and MdPHT5 has three. Our overview of this gene family focused on structure, chromosomal distribution and localization, phylogenies, and motifs. These genes displayed differential expression patterns in various tissues. For example, expression was high for MdPHT1;12, MdPHT3;6, and MdPHT3;7 in the roots, and was also increased in response to low-phosphorus conditions. In contrast, MdPHT4;1, MdPHT4;4, and MdPHT4;10 were expressed only in the leaves while transcript levels of MdPHT1;4, MdPHT1;12, and MdPHT5;3 were highest in flowers. In general, these 37 genes were regulated significantly in either roots or leaves in response to the imposition of phosphorus and/or drought stress. The results suggest that members of the PHT family function in plant adaptations to adverse growing environments. Our study will lay a foundation for better understanding the PHT family evolution and exploring genes of interest for genetic improvement in apple.

Keywords: apple; drought; expression; gene family; phosphate transporters; phosphorus stress.

Figure 1

Positions of PHT gene family members on apple chromosomes.

Figure 2

(A) Phylogenetic analysis. Different colors and shapes indicate genes within individual clusters. (B) Schematic diagram of exon/intron structures for PHT genes in apple. Diagram was made via Gene Structure Display Server, applying method for coding sequences and corresponding genome sequences. Yellow boxes and gray lines represent exons and introns, respectively.

Figure 3

Phylogenetic analyses of PHT genes in Arabidopsis thaliana (At), Oryza sativa (Os), Malus domestica (Md), and Populus trichocarpa (Ptr). Unrooted phylogenetic tree was constructed by NJ method, using MEGA6.0 program.

Figure 4

Protein sequences identified from MdPHT family and aligned with MEME software.

Figure 5

Quantitative real-time PCR analysis of selected apple MdPHT1 genes (A), MdPHT2 genes (B), MdPHT3 genes (C), MdPHT4 genes (D), MdPHT5 genes (E) expressed in roots, stems, leaves, shoot tips, flowers, young fruits and mature fruits from Malus hupehensis var. pingyiensis. Data were normalized to level of apple Actin expression. Value for each sample is mean of 3 replicates. Vertical bars indicate standard deviation.

Figure 6

The expression profiles of PHT genes in leaves of Malus hupehensis var. pingyiensis. Heatmap shows PHT gene expression across after 15 d of phosphorus treatment, samples were taken from leaves exposed to low-P (LLP), high-P (LHP), drought (LD), drought with low-P (LDLP), or drought with high-P (LDHP) conditions. All transcript levels were normalized to their respective corresponding levels in non-stressed control (LCK).

Figure 7

The expression profiles of PHT genes in roots of Malus hupehensis var. pingyiensis. Heatmap shows PHT gene expression across after 15 d of phosphorus treatment, samples were taken from roots exposed to low-P (RLP), high-P (RHP), drought (RD), drought with low-P (RDLP), or drought with high-P (RDHP) conditions. All transcript levels were normalized to their respective corresponding levels in non-stressed control (RCK).