Cellular Heterogeneity and Developmental Dynamics of Aril in Papaya

The papaya aril is a specialized seed appendage that has been reported to contain germination-inhibiting substances and usually requires removal before seed germination, thereby limiting breeding efficiency. However, the cellular origin and candidate molecular regulators of papaya aril development remain poorly understood. To investigate the early developmental process and candidate regulatory genes of the papaya aril, we combined histological analysis, bulk RNA-seq, and single-cell RNA-seq. Histological observations suggested that aril differentiation begins around 10 days after pollination (DAP) in the funiculus region. Based on this initiation stage, bulk RNA-seq profiling of seeds at 5, 10, and 15 DAP identified genes with initiation-stage-specific expression and prioritized candidate genes potentially related to seed appendage development, including CpRING-like, CpMBR2, and CpNDR8. Single-cell RNA-seq of seeds at 10 and 15 DAP annotated a putative aril cell population and reconstructed its developmental trajectory, revealing five trajectory-associated genes: CpATJ3, CpDYL1, CpGRP-like, CpHIRD11, and CpERD15. Integrative analysis of bulk and single-cell transcriptomic datasets further identified three candidate genes potentially involved in aril development: CpFER3, CpUVI4, and CpCEP1. These findings support the funiculus region as the likely anatomical origin of the papaya aril and provide candidate genes for future functional validation.