The evolutionary trajectories and gene regulatory roles of nuclear-integrated plastid DNA: clues for enhancing environmental adaptation in Caryophyllales

Abstract

Environmental stimuli can induce the transfer of chloroplast DNA to the nuclear genome, resulting in nuclear-integrated plastid DNAs (NUPTs). However, their role in plant adaptability remains unclear. Species within the Caryophyllales order, known for their adaptation to extreme environments, provide an ideal model for studying the evolutionary dynamics and functions of NUPTs. In this study, we analyzed NUPTs in 24 Caryophyllales species to investigate their evolution and regulatory roles in gene expression, particularly in response to environmental stimuli. We found significant interspecies variation in NUPT abundance, ranging from 566 insertions in Amaranthus cruentus to 3585 in Beta vulgaris, with sizes spanning from 100 bp to over 100 kb. Approximately 62% of NUPTs were inserted within the last 20 million years, while some species exhibit insertion peaks dating back 49 million years. NUPT presence/absence polymorphisms in six related species suggest that NUPT insertions and deletions are dynamic processes influenced by phylogeny. NUPTs predominantly integrate into intergenic regions but also insert into genes and promoters, with certain regions acting as hotspots. Notably, NUPTs introduce numerous environmental-responsive cis-acting elements in promoter regions. Genes with NUPT insertions in their promoters are significantly enriched for functions related to environmental response. Further luciferase assays in Spinacia oleracea demonstrated that NUPT insertions can regulate the expression of genes related to environmental responses, indicating their potential role in adaptive evolution. Overall, our study provides insights into NUPT evolution and their influence on gene function and plant adaptability to environmental stimuli.