Soybean Cyst Nematode-Resistant Protein AATRhg1 Affects Amino Acid Homeostasis and Betalain Accumulation

Amino acid transporters play crucial roles in plant nitrogen metabolism but also defense responses. AATRhg1, an apparent amino acid transporter encoded by Glyma.18g022400 (Rhg1-GmAAT) at the soybean Rhg1 locus, contributes to resistance to soybean cyst nematode (SCN), although the in planta function of AATRhg1 remains elusive. In this study we discovered that overexpression of Rhg1-GmAAT in soybean roots enhances the betalain pigment synthesis driven by a RUBY transgene cassette, potentially through its transporter activity affecting tyrosine level and amino acid homeostasis. Silencing Rhg1-GmAAT also moderately increased betalain accumulation while co-overexpression of Rhg1-GmAAT and GmRBOHG (encoding an AATRhg1-interacting NADPH oxidase) blocked the betalain phenotype, indicating a complex role of AATRhg1 in regulating cellular metabolism. Soybean AATRhg1 did not show a betalain accumulation phenotype when co-overexpressed with RUBY in Nicotiana benthamiana leaves, suggesting that soybean AATRhg1 functions differently in N. benthamiana. In soybean, the expression of AATRhg1 proteins mutated at conserved residues D122A or Y268L mitigated or enhanced the betalain phenotypes, respectively, suggesting that these residues are important for AATRhg1 function. This study advances our understanding of AATRhg1 while presenting a novel strategy for enhancing betalain biosynthesis by modulating transport and homeostasis of amino acids.