LIKE EARLY STARVATION is involved in the regulation of starch initiation in potato (Solanum tuberosum cv. Désirée) tubers.

Starch initiation has been intensively investigated. In tubers of Solanum tuberosum, it is controlled by a heteromultimeric isoamylase complex composed of ISA1 and ISA2. Moreover, two non-catalytic starch binding proteins (i.e., ESV1 and LESV) regulate leaf starch metabolism in Arabidopsis. Both proteins interact with starch glucans through a C-terminal antiparallel ß-sheet domain and differ by the presence of a 115 amino acid N-terminal overhang in LESV. Here we report the CRIPSR/Cas9 inactivation of LESV or ESV1 in potato and the characterization of the respective tuber starches. Starch from esv1 mutants was unaffected whereas granule diameter was severely reduced in tubers of lesv plants. This was negatively correlated to the number of granules, coherent with an altered starch initiation phenotype. Strikingly, scanning electron microscopy of purified starches revealed that lesv plants phenocopy isa1 and isa2 antisense lines described 20 years ago. Here, we confirmed that LESV interacts with ISA1 in potato using yeast two-hybrid assays, as demonstrated in rice, providing molecular evidence for their functional association in potato. This interaction supports the hypothesis that LESV acts in concert with the ISA1/ISA2 complex, likely regulating the early steps of glucan organization required for proper starch granule formation.