SaRac1 , a small GTPase, plays an essential role in haustorium development in the root parasite sandalwood ( Santalum album )

Abstract Indian sandalwood (Santalum album) is an economically important facultative parasite that develops a specialized multicellular organ, the haustorium, to absorb water and nutrients from its hosts. To elucidate the molecular mechanisms underlying haustorium development, we conducted a transcriptome analysis across six S. album tissues. We found that SaRac1, encoding a functional small GTPase, is specifically expressed in the haustorium. We employed host-induced gene silencing (HIGS) by generating transgenic poplar (Populus alba × P. glandulosa) hosts that express hairpin RNAs to target and downregulate SaRac1 in the parasite. Santalum album grown with SaRac1 RNAi transgenic host plants exhibited significantly suppressed haustorium development compared with those grown with wild-type or empty-vector controls. Mechanistically, SaRac1 interacts with SaRbohA, and this interaction synergistically enhances ROS production. Exogenous H₂O₂ application significantly upregulated key haustorium formation-related genes. In contrast, the Rboh inhibitor diphenyliodonium chloride (DPI) suppressed the expression of SaYUCCA and SaSBT in S. album grown with wild-type and empty-vector control hosts, thereby reducing haustorium formation. In S. album plants grown with RNAi hosts, SaSBT and SaEXPA were also downregulated by DPI application. Our findings identify a crucial mechanism whereby SaRac1 promotes haustorium formation by modulating ROS signaling and provide novel insights into the molecular physiology of plant parasitism.