Full-Length Transcriptome Analysis of Alternative Splicing and Polyadenylation in the Molecular Regulation of Labor Division in Apis cerana cerana

Honeybees are vital pollinators with functional differentiation as a key survival strategy. The Chinese honeybee (<i>Apis cerana cerana</i>) exhibits exceptional nectar foraging in complex terrains, yet how alternative splicing (AS) and polyadenylation (APA) regulate its labor division remains unclear. Here, we applied PacBio full-length transcriptome sequencing to annotate worker bee transcriptomes across three developmental stages (<i>Ac</i>3d, <i>Ac</i>10d, <i>Ac</i>21d), calibrating the third-generation sequencing data with second-generation sequencing to enhance transcriptome annotation accuracy. We identified 17,961 isoforms and 1922 APA genes, finding that alternative first exon was the major type of AS, while APA enhances transcriptomic diversity via dual polyadenylation sites in most genes. Functional analyses revealed AS enrichment in growth signaling (<i>Vg6, CYP15A1</i>) and immune pathways (<i>PTPRR</i>), whereas APA regulated growth signaling (<i>Vg6</i>), energy metabolism (<i>Acsl1, AcceFE</i>), and oxidative stress (<i>PTPRR, PPO2</i>). Validation by PCR and 3'RACE confirmed stage-specific AS/APA events in key genes. These findings significantly enhance the <i>A. cerana cerana</i> reference genome annotation and provide valuable insights into the mechanisms of AS and APA regulation underlying honeybee development and functional transitions.