Professor University of Florida Lake Alfred, FL, USA
Abstract Text: Citrus Huanglongbing (HLB) caused by Candidatus Liberace asiaticus (CLas) is the most devastating citrus disease worldwide. CLas resides in the phloem and is naturally vectored by Asian citrus psyllids. No commercial citrus cultivars are resistant to HLB. HLB has been reported to be an immune-mediated disease. CLas triggers systemic and chronic immune responses, including reactive oxygen species (ROS) production and callose deposition, leading to phloem cell death and HLB symptoms. We hypothesize that enhancing the expression level of antioxidant enzymes mitigates ROS levels, thus increasing citrus tolerance against HLB. Here, we aim to increase the protein expression level of antioxidant enzymes, including Cu/Zn superoxide dismutase (Cu/Zn SOD), ascorbate peroxidase (APX), and glutathione peroxidase (GPX) by editing the upstream open reading frames (uORFs) regions. uORFs are short protein-coding elements located in the 5’UTR region of the downstream primary ORF (pORF). Previous studies have demonstrated that uORFs normally repress the translation level of pORF mRNAs. Thus, deletion of uORFs usually leads to increased protein expression of pORFs. We conducted genome editing of citrus by transforming embryogenic protoplasts of citrus using the ribonucleoprotein complex (RNP) consisting of Cas12a and gRNAs targeting the uORFs of antioxidant enzyme genes. This study provides a potential strategy for developing transgene-freeHLB-resistant/tolerantcitrus varieties via genome editing, offering an efficient and sustainable solution against HLB for citrus growers.
