Citrus Huanglongbing (HLB), caused by the phloem-restricted bacterium Candidatus Liberibacter asiaticus (CLas), is a devastating disease threatening global citrus production. CLas infection triggers excessive accumulation of phloem proteins (PPs) that obstruct sieve pores, a dual-edged process potentially restricting pathogen spread while impairing phloem transport. Despite its pathophysiological significance, systematic identification and functional characterization of PPs in citrus, particularly their roles in CLas defense, remain unclear. Here, we performed a genome-wide analysis of the PP2 gene family in the HLB-susceptible sweet orange (Citrus sinensis) and identified 26 CsPP2 genes. Phylogenetic and structural analyses uncovered evolutionary divergence and regulatory complexity among CsPP2 family members. Using promoter-driven GUS gene expression assays in transgenic hairy roots, we identified three phloem-specific paralogs, CsPP2-3, CsPP2-5, and CsPP2-18, and delineated core regulatory regions conferring tissue specificity. Overexpression of each gene significantly enhanced phloem protein deposition. Notably, although virus-induced silencing of individual CsPP2s did not affect resistance to Xanthomonas citri subsp. citri, overexpression of any of the three genes substantially enhanced resistance against this apoplastic pathogen, demonstrating functional redundancy. However, the three paralogs exhibited marked functional divergence in response to CLas: CsPP2-3 and CsPP2-18 conferred enhanced resistance, whereas CsPP2-5 increased susceptibility. Distinct defense-related gene expression profiles further supported their specialized immune roles. Our study provides the first systematic identification of PP2 genes in citrus and reveals the functional differentiation of CsPP2-3/5/18 as key regulators of phloem-mediated defense. These findings provide crucial insights into phloem defense regulatory networks and identify novel genetic targets for HLB resistance breeding.

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