ABA-responsive NnABI4 coordinates transitory and storage starch accumulation by activating NnSUS3 and NnSBEII in lotus

Abstract

The diurnal synthesis and degradation of transitory starch are critical for photosynthetic carbon fixation and allocation to sink organs. Although abscisic acid (ABA) is known to influence carbon partitioning, its regulatory role in starch metabolism and yield formation in lotus remains unclear. Here, we show that ABSCISIC ACID INSENSITIVE 4 (NnABI4), an ABA-responsive transcription factor, positively regulates both transitory and storage starch biosynthesis in lotus. Transient overexpression and silencing assays confirmed that NnABI4 promotes starch accumulation while reducing the amylose/amylopectin ratio. To substantiate these findings, we generated nine independent transgenic potato lines. These lines displayed enhanced leaf growth, increased transitory starch content in leaves, and substantially higher tuber yield, starch granule number, and amylopectin content, as well as modified tuber physicochemical properties. Mechanistically, NnABI4 is transcriptionally induced by ABA and directly binds to the CACCG motif in the promoters of sucrose synthase 3 (NnSUS3) and starch-branching enzyme II (NnSBEII), activating their expression. Complementation assays revealed that overexpression of either NnSUS3 or NnSBEII partially rescued the starch-deficient phenotype in NnABI4-silenced rhizomes. In leaves, however, NnSUS3 overexpression increased sucrose levels without restoring starch content, whereas NnSBEII overexpression elevated total starch and amylopectin while reducing sucrose, indicating distinct, tissue-dependent functions of these two downstream genes. Collectively, our results reveal a dual molecular mechanism through which ABA–NnABI4 signaling directly modulates NnSUS3 and NnSBEII in a branch- and organ-specific manner. This work provides a theoretical basis and identifies candidate genetic targets for improving yield and quality traits in lotus and other root and tuber crops through molecular breeding.