The Transcription Factor LpWRKY65 Enhances Embryogenic Capacity through ROS Scavenging during Somatic Embryogenesis of Larch

Abstract

Somatic embryogenesis is a powerful system for studying embryo development and scaling up the production of elite genetic material. Somatic embryogenesis has been well established in Larix principis-rupprechtii, a Chinese larch species dominant in the world's largest man-made forest. However, genotype-dependent embryogenic variations hinder large-scale forestry, and the molecular mechanisms remain unclear. Here, we constructed stage-specific developmental transcriptomes of the somatic embryogenesis process using two lines with contrasting embryogenic capacities. Clustering and co-expression analyses identified LpWRKY65 as a central hub gene highly expressed in early somatic embryogenesis stages and with significantly higher expression in the high-embryogenic-capacity cell line (HEL) compared to the low-embryogenic-capacity cell line (LEL). Overexpressing LpWRKY65 significantly increased somatic embryo yield and quality. DAP-seq and RNA-seq were combined to identify a set of target genes downstream of and responsive to LpWRKY65, particularly including genes involved in reactive oxygen species (ROS) scavenging. We identified LpHmgB10 as a critical downstream regulator of LpWRKY65. LpWRKY65 directly binds to the W-box in the promoter of LpHmgB10, markedly enhancing its transcriptional activity. ROS profiling further demonstrated that overexpression of LpWRKY65 or LpHmgB10 enhances ROS scavenging and promotes a stable redox environment, which is crucial for improving embryogenic capacity. These findings suggest that LpWRKY65 regulates the cellular redox environment to promote embryogenic differentiation and somatic embryo development, advancing somatic embryogenesis research in conifers.