High-efficiency TALEN-based gene editing produces disease-resistant rice

Journal Article / 2012

Li, Ting; Liu, Bo; Spalding, Martin H.; Weeks, Donald P.; Yang, Bing

Transcription activator–like (TAL) effectors of Xanthomonas oryzae pv. oryzae (Xoo) contribute to pathogen virulence by transcriptionally activating specific rice disease-susceptibility (S) genes1,2. TAL effector nucleases (TALENs)—fusion proteins derived from the DNA recognition repeats of native or customized TAL effectors and the DNA cleavage domains of FokI3–5—have been used to create site-specific gene modifications in plant cells6,7, yeast8, animals9–12 and even human pluripotent cells13. Here, we exploit TALEN technology to edit a specific S gene in rice to thwart the virulence strategy of X. oryzae and thereby engineer heritable genome modifications for resistance to bacterial blight, a devastating disease in a crop that feeds half of the world’s population. We targeted the rice bacterial blight susceptibility gene Os11N3 (also called OsSWEET14) for TALEN-based disruption. This rice gene encodes a member of the SWEET sucrose-efflux transporter family and is hijacked by X. oryzae pv. oryzae, using its endogenous TAL effectors AvrXa7 or PthXo3, to activate the gene and thus divert sugars from the plant cell so as to satisfy the pathogen’s nutritional needs and enhance its persistence2,14. The Os11N3 promoter contains an effector-binding element (EBE) for AvrXa7, overlapping with another EBE for PthXo3 and with the TATA box (Fig. 1a and Supplementary Fig. 1). We deployed two pairs of designer TALENs (pair 1 and pair 2) independently to induce mutations in these overlapping EBEs of the Os11N3 promoter and thus to interfere with the virulence function of AvrXa7 and PthXo3, but not the developmental function of Os11N3 (Supplementary Fig. 1 and Supplementary Note). The TALE repetitive regions used for nuclease fusions included the native AvrXa7 and three designer TALE repetitive regions custom synthesized using a modular assembly method8. Each designer TALEN contained 24 repeat units for recognition of a specific set of 24 contiguous nucleotides at the target sites (Supplementary Fig. 1).