Genome-Wide Identification of the ARF Gene Family and ARF3 Target Genes Regulating Ovary Initiation in Hazel via ChIP Sequencing

Hazel (Corylus spp.) is an economically important nut species with a unique biological characteristic of ovary differentiation and development initiating from the ovary primordium after pollination. Auxin participates in ovary initiation and has an essential impact on hazel fruit yield and quality. The regulation of auxin in ovary development is thought to be related to auxin response factors (ARFs); however, its detailed regulatory mechanism remains unclear. The spatiotemporal expression pattern of C. heterophylla ARF3 (ChARF3) was accessed via ARF gene family member identification and expression abundance analysis as well as immunohistochemistry. ChARF3 target genes were identified via chromatin immunoprecipitation followed by next-generation sequencing (ChIP-Seq). In total, 14 ChARF members containing at least B3 and Auxin_resp domains were found to be distributed on 9 of 11 chromosomes, and the protein molecular weights were predicted to range from 70.93–139.22 kD. Among eight differentially expressed ChARFs, ChARF3 showed the most significant differences over four ovary developmental stages. Immunohistochemical analysis revealed that ChARF3 was expressed in the ovary primordium and funiculus, integument, endosperm, radicle, and cotyledon indicating its potential regulatory roles in ovary differentiation and development. In total, 3,167 ChARF3 target genes were identified through ChIP-Seq in four ovary developmental stages and were significantly enriched in the biosynthesis of secondary metabolites (ko01110), phenylpropanoid biosynthesis (ko00940), and phytohormone signal transduction (ko04075). ChARF3 was hypothesized to be involved in the regulation of auxin-induced genes and the transcription factors MADS, AP2/ERF, TCP, FT, and LFY. These results suggest that ChARF3 may regulate ovary initiation and ovule development by mediating genes related to auxin biosynthesis and transport, cell division and proliferation, and flower and fruit development. This study provides new insights into the molecular mechanism of hazel yield formation.