A NAC-centered transcriptional network as a core regulatory module controlling the onset of grape berry ripening

Abstract

The onset of fruit ripening represents a major developmental transition characterized by extensive transcriptional and metabolic reprogramming that determines fruit texture, color, flavor, and aroma. In grapevine (Vitis vinifera L.), rising temperatures due to global warming advance the onset of ripening and reshape berry physiology, underscoring the need to resolve the transcriptional regulatory network controlling this transition. NAC transcription factors constitute a prominent component of these regulatory circuits, with VviNAC33, VviNAC60, and VviNAC03 implied in cooperative regulation of ripening-related processes, and VviNAC61 linked to later stages of ripening. Moreover, numerous additional NAC genes are specifically induced during grape berry development and ripening, yet their functional roles and network connectivity remain largely unexplored. Here, we investigate twelve berry-specific NAC genes as candidate master regulators driving large-scale transcriptome reprogramming during grapevine development. By integrating gene regulatory network inference based on co-expression, homologous overexpression coupled with transcriptomic profiling, cistrome mapping by DNAaffinity purification sequencing (DAP-seq), and complementary molecular approaches, we dissect intra-family regulatory interactions and the ripening-related processes they control. Based on temporal expression profiles in early-and late-ripening cultivars, together with transient overexpression in Fragaria vesca and Fragaria × ananassa fruit, VviNAC60 and VviNAC18 emerge as the earliest NAC transcription factors involved in the initiation of ripening. Collectively, our results define a NAC-centered transcriptional network as a core regulatory module controlling the onset of grape berry ripening.