Integrated genomic, transcriptomic, and proteomic analyses prioritize cadmium-responsive class III peroxidases in Salix variegata

Abstract

Class III peroxidases (PRXs) link apoplastic redox regulation with cell-wall remodeling, yet their coordinated transcript- and protein-level responses to cadmium (Cd) remain poorly resolved in woody plants. We identified 80 high-confidence class III PRX genes in Salix variegata and characterized their genomic organization, evolutionary relationships, protein properties, and predicted regulatory features. Root RNA-seq and data-independent acquisition proteomic datasets from plants exposed to 0 or 100 μM CdCl2 for 10 d were then integrated to prioritize members supported across molecular layers. Segmental and dispersed duplication were the predominant expansion modes, and most SvaPRX proteins showed sequence features compatible with extracellular localization. RNA-seq identified 42 upregulated and two downregulated SvaPRX genes. The DIA dataset comprised 22,883 peptides and 6,068 proteins, with 4,849 proteins quantified in all six samples; biological replicates showed high within-treatment correlations and separation of CK and Cd100 in principal component analysis. Forty SvaPRX proteins were quantified, including 11 upregulated and six downregulated proteins. Cross-layer integration reduced the broad transcript-responsive set to seven concordantly upregulated candidates: SvPRX12, SvPRX23, SvPRX41, SvPRX47, SvPRX61, SvPRX62, and SvPRX66. Targeted qRT-PCR provided additional directional support for Cd-induced expression of SvPRX61 and SvPRX62, with response directions consistent across RNA-seq, proteomic, and qRT-PCR datasets. These results establish a reproducible genome-to-proteome framework for prioritizing Cd-responsive class III peroxidases and identify focused candidates for subsequent biochemical and genetic investigation in woody plants.