Pigeonpea (Cajanus cajan) is an important legume food crop and plays a crucial role in a secure food supply in many developing countries. Several previous studies have suggested that pigeonpea has great potential for phosphorus (P) deficiency tolerance, but little is known about the underlying mechanism. In this study, the physiological and molecular responses of pigeonpea roots to phosphate (Pi) starvation were investigated through integrating phenotypic, genomic, transcriptomic, metabolomic, and lipidomic analyses. The results showed that low-Pi treatment increased total root length, root surface area, and root acid phosphatase activity, and promoted the secretion of organic acids (e.g. citric acids, piscidic acids, and protocatechuic acids) and the degradation of phospholipids and other P-containing metabolites in the roots of pigeonpea. Consistent with the morphological, physiological, and biochemical changes, a large number of genes involved in these Pi-starvation responses were significantly upregulated in Pi-deficient pigeonpea roots. Among these Pi-starvation response genes upregulated by low-Pi treatment, four gene families were expanded through recent tandem duplication in the pigeonpea genome, namely phosphate transporter 1 (PHT1), phosphoethanolamine/phosphocholine phosphatase (PECP), fasciclin-like arabinogalactan protein (FLA), and glutamate decarboxylase (GAD). These gene families may be associated with Pi uptake from the soil, phospholipid recycling, root morphological remodeling, and regulation of organic acid exudation. Taken together, our results suggest that pigeonpea employs complex Pi-starvation responses to strengthen P acquisition and utilization during low-Pi stress. This study provides new insights into the genome evolution and P deficiency adaptation mechanism of pigeonpea.

Senna tora is one of the homologous crops used as a medicinal food containing an abundance of anthraquinones. Type III polyketide synthases (PKSs) are key enzymes that catalyze polyketide formation; in particular, the chalcone synthase-like (CHS-L) genes are involved in anthraquinone production. Tandem duplication is a fundamental mechanism for gene family expansion. However, the analysis of the tandem duplicated genes (TDGs) and the identification and characterization of PKSs have not been reported for S. tora. Herein, we identified 3087 TDGs in the S. tora genome; the synonymous substitution rates (Ks) analysis indicated that the TDGs had recently undergone duplication. The Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis showed that the type III PKSs were the most enriched TDGs involved in the biosynthesis of the secondary metabolite pathways, as evidenced by 14 tandem duplicated CHS-L genes. Subsequently, we identified 30 type III PKSs with complete sequences in the S. tora genome. Based on the phylogenetic analysis, the type III PKSs were classified into three groups. The protein conserved motifs and key active residues showed similar patterns in the same group. The transcriptome analysis showed that the chalcone synthase (CHS) genes were more highly expressed in the leaves than in the seeds in S. tora. The transcriptome and qRT-PCR analysis showed that the CHS-L genes had a higher expression in the seeds than in other tissues, particularly seven tandem duplicated CHS-L2/3/5/6/9/10/13 genes. The key active-site residues and three-dimensional models of the CHS-L2/3/5/6/9/10/13 proteins showed slight variation. These results indicated that the rich anthraquinones in S. tora seeds might be ascribed to the PKSs’ expansion from tandem duplication, and the seven key CHS-L2/3/5/6/9/10/13 genes provide candidate genes for further research. Our study provides an important basis for further research on the regulation of anthraquinones’ biosynthesis in S. tora.