Evolutionary origin of a new gene drives root growth in one desert poplar

Populus euphratica develops an extensive root system that supports survival in arid environments. We identified PeuPLDζ4, a new gene arising from tandem duplication in P. euphratica, and evaluated its effects on root growth and drought-related performance. Phylogenetic and promoter-reporter analyses, enzyme assays, and subcellular localization characterized PeuPLDζ4 divergence. Its effects were evaluated in transgenic Arabidopsis thaliana, P. alba var. pyramidalis, wheat, and rice. Phosphatidic acid (PA) and hormone measurements, reporter imaging, transcriptomics, protein-lipid overlay, dual-luciferase, and electrophoretic mobility shift assays investigated the mechanism. Compared with its paralog PeuPLDζ3, PeuPLDζ4 showed drought-inducible root expression, higher enzymatic activity, and additional nuclear-associated localization. PeuPLDζ4 overexpression promoted root growth across species and was associated with increased PA accumulation in root-tip tissues, elevated auxin (IAA) and cytokinin (CK) contents, enhanced IAA and CK reporter responses, and an increased IAA : CK ratio. NAC097 was identified as a PA-associated candidate regulator that bound and activated the LONELY GUY 2 (LOG2) and IIOPENTENYLTRANSFERASE 4 (IPT4) promoters. These findings indicate that functional divergence of a newly duplicated PLDζ gene contributed to root-related adaptation in P. euphratica. The PA-associated NAC097-LOG2/IPT4 branch provides a candidate link between lipid signaling and CK regulation, while PeuPLDζ4 may offer a resource for improving drought-related traits in crops.