The miR395b–ABI5 module regulates amylopectin branching and biosynthesis and affects lotus root quality

Starch is an important carbohydrate in lotus (Nelumbo nucifera Gaertn.) roots, which are edible rhizomes, and starch biosynthesis and metabolism strongly influence lotus root yield and quality. Plant microRNAs (miRNAs) have a significant effect on crop yield, quality, and starch biosynthesis, but the molecular mechanism by which miRNAs regulate starch biosynthesis is unknown. In this study, miR395b expression levels showed significant differences in lotus germplasms with different starch contents. Overexpressing MIR395b reduced amylopectin levels and medium- and long-branched chain percentages in lotus roots, decreasing total starch accumulation. Inhibiting miR395b function (using short tandem target mimics) or overexpressing ABA Insensitive 5 (NnABI5) increased amylopectin content and medium- and long-branched chain proportions in lotus roots, significantly enhancing total starch content. miR395b specifically targeted and repressed the expression of the transcription factor NnABI5. Furthermore, NnABI5 promoted amylopectin biosynthesis and medium- and long-branched chain proportions in lotus roots by binding to the promoter region of the amylopectin biosynthesis-related genes starch synthetase 2 (NnSS2) and starch branching enzyme II (NnSBEII) and activating their expression. Our study reveals the critical role of the miR395b-NnABI5 module in starch biosynthesis in lotus roots, providing a theoretical foundation for lotus root quality and yield improvement.