Polyploidization-Driven Functional Innovation of AGPase Small Subunit Gene APS1 Regulates Starch Biosynthesis in Banana (Musa acuminata)

Starch biosynthesis is a fundamental process influencing yield and fruit quality in banana, with ADP-glucose pyrophosphorylase (AGPase) serving as the rate-limiting enzyme catalyzing sucrose conversion into starch. However, the mechanisms underlying functional differentiation of AGPase family genes following polyploidization remain largely unexplored. In this study, eight AGPase genes, including large (MaAPL) and small subunit (MaAPS) members, were identified from the banana (Musa acuminata) genome, all harboring the conserved ADP-glucose pyrophosphorylase domain. Phylogenetic analysis traced their evolutionary origin to the ancient moss Physcomitrella patens, with polyploidization identified as the primary driver of gene family expansion. These genes exhibit conserved codon usage bias and have undergone strong purifying selection. Among them, MaAPS1 displayed distinct functional differentiation, increased intron number, enriched promoter cis-elements, and significantly elevated expression—features likely contributing to its adaptation for enhanced starch accumulation in fruit. Furthermore, the MaAPS1 protein was predominately localized in the chloroplast. Functional validation supported its regulatory involvement: transient silencing in banana fruit reduced starch content, while transient overexpression in banana fruit increased starch levels. Co-expression and molecular docking analyses revealed that transcription factors ERF1, C3H1, bZIP1, and bZIP3 may interact with the MaAPS1 promoter, indicating a multifactorial regulatory network. Overall, this study provides insights into polyploidy-driven functional innovation and transcriptional regulation of MaAPS1 in banana starch biosynthesis, providing valuable molecular targets for genetic improvement of yield and fruit quality.