A natural mutation of the <i>NST1</i> gene arrests secondary cell wall biosynthesis in the seed coat of a hull-less pumpkin accession

Abstract

Hull-less pumpkins (<i>Cucurbita pepo</i> L.) are naturally occurring novel variants known as oilseed or naked-seeded pumpkins, and are characterized by the absence of a normal lignified seed coat. Due to a specialized seed coat structure, these variants serve as a good model for studying seed coat formation and simplify the processing of pumpkin seeds. However, causal genes for this hull-less trait still remain unknown. Here, by bulked segregant analysis and fine mapping, we found that mutation of a single gene, <i>NAC SECONDARY WALL THICKENING PROMOTING FACTOR 1</i> (<i>NST1</i>), accounts for the hull-less trait. A 14-bp sequence insertion in the <i>CpNST1</i> gene causes premature termination of <i>CpNST1</i> translation, leading to lack of secondary cell wall (SCW) biosynthesis in hull-less seed coats. <i>In situ</i> hybridization analysis provided further evidence for the role of <i>CpNST1</i> in pumpkin seed coat SCW biosynthesis. Interestingly, through secondary cell wall compositional analysis, we found that the main SCW components differed among cell layers in the seed coat. RNA-seq analysis indicated an upstream role of CpNST1 in the SCW biosynthesis network. Collectively, our findings provide mechanistic insight into seed coat SCW biosynthesis, and a target gene for breeders to introduce this hull-less trait for commercial exploitation.