PtrPSK2a is Required for Cambial Development and Xylem Formation during Secondary Growth in Poplar

Phytosulfokines (PSKs) are sulfated peptide hormones involved in plant growth, development, and stress responses. However, direct genetic evidence for the involvement of PSK peptide in cambium development and secondary vascular development in woody plants remains largely lacking. Here, we identified nine PSK family members in Populus trichocarpa. Phylogenetic, gene structure, conserved motif, and local synteny analyses indicated that the poplar PSK family is evolutionarily conserved, and some members may have arisen from the whole-genome duplication (WGD) event. Subsequently, we generated overexpression and CRISPR/Cas9-mediated knockout lines of PtrPSK2a in hybrid poplar 717. PtrPSK2a is preferentially expressed in stems, particularly in the cambium and developing xylem. Both gain-of-function and loss-of-function of PtrPSK2a exhibited growth defects, characterized by reduced plant height and stem diameter, with more severe overall growth defects in OE lines. Consistently, histological analysis demonstrated that cambium activity and secondary xylem development were significantly inhibited in both transgenic lines, as reflected by fewer cambium cell layers, narrower xylem, and fewer vessels in young stems. In addition, genes associated with cambial development, vessel differentiation, and secondary cell wall formation were broadly downregulated, accompanied by altered expression of several hormone-related marker genes in transgenic lines. Together, these findings demonstrate that PtrPSK2a is essential for cambial activity and xylem development during secondary growth in poplar.