Bacterial blight of rice is an important disease in Asia and Africa. The pathogen, Xanthomonas oryzae pv. oryzae (Xoo), secretes one or more of six known transcription-activator-like effectors ...(TALes) that bind specific promoter sequences and induce, at minimum, one of the three host sucrose transporter genes SWEET11, SWEET13 and SWEET14, the expression of which is required for disease susceptibility. We used CRISPR-Cas9-mediated genome editing to introduce mutations in all three SWEET gene promoters. Editing was further informed by sequence analyses of TALe genes in 63 Xoo strains, which revealed multiple TALe variants for SWEET13 alleles. Mutations were also created in SWEET14, which is also targeted by two TALes from an African Xoo lineage. A total of five promoter mutations were simultaneously introduced into the rice line Kitaake and the elite mega varieties IR64 and Ciherang-Sub1. Paddy trials showed that genome-edited SWEET promoters endow rice lines with robust, broad-spectrum resistance.
Non-coding small RNAs (sRNA) act as mediators of gene silencing and regulate plant growth, development and stress responses. Early insights into plant sRNAs established a role in antiviral defense ...and they are now extensively studied across plant-microbe interactions. Here, sRNA sequencing discovered a class of sRNA in rice (Oryza sativa) specifically associated with foliar diseases caused by Xanthomonas oryzae bacteria. Xanthomonas-induced small RNAs (xisRNAs) loci were distinctively upregulated in response to diverse virulent strains at an early stage of infection producing a single duplex of 20-22 nt sRNAs. xisRNAs production was dependent on the Type III secretion system, a major bacterial virulence factor for host colonization. xisRNA loci overlap with annotated transcripts sequences, with about half of them encoding protein kinase domain proteins. A number of the corresponding rice cis-genes have documented functions in immune signaling and xisRNA loci predominantly coincide with the coding sequence of a conserved kinase motif. xisRNAs exhibit features of small interfering RNAs and their biosynthesis depend on canonical components OsDCL1 and OsHEN1. xisRNA induction possibly mediates post-transcriptional gene silencing but they do not broadly suppress cis-genes expression on the basis of mRNA-seq data. Overall, our results identify a group of unusual sRNAs with a potential role in plant-microbe interactions.
Bacterial leaf blight (BB) of rice, caused by
pv.
(
), threatens global food security and the livelihood of small-scale rice producers. Analyses of
collections from Asia, Africa and the Americas ...demonstrated complete continental segregation, despite robust global rice trade. Here, we report unprecedented BB outbreaks in Tanzania. The causative strains, unlike endemic African
, carry Asian-type TAL effectors targeting the sucrose transporter
and iTALes suppressing
. Phylogenomics clustered these strains with
from Southern-China. African rice varieties do not carry effective resistance. To protect African rice production against this emerging threat, we developed a hybrid CRISPR-Cas9/Cpf1 system to edit all known TALe-binding elements in three
promoters of the East African elite variety Komboka. The edited lines show broad-spectrum resistance against Asian and African strains of
, including strains recently discovered in Tanzania. The strategy could help to protect global rice crops from BB pandemics.
pv.
(
) strains that cause bacterial leaf blight (BLB) limit rice (
) production and require breeding more resistant varieties. Transcription activator-like effectors (TALEs) activate transcription ...to promote leaf colonization by binding to specific plant host DNA sequences termed effector binding elements (EBEs).
major TALEs universally target susceptibility genes of the SWEET transporter family. TALE-unresponsive alleles of clade III
susceptibility gene promoter created with genome editing confer broad resistance on Asian
strains. African
strains rely primarily on the major TALE TalC, which targets
. Although the virulence of a
mutant strain is severely impaired, abrogating
induction with genome editing does not confer equivalent resistance on African
. To address this contradiction, we postulated the existence of a TalC target susceptibility gene redundant with
. Bioinformatics analysis identified a rice locus named ATAC composed of the
(
) gene and a putative lncRNA that are shown to be bidirectionally upregulated in a TalC-dependent fashion. Gain-of-function approaches with designer TALEs inducing ATAC sequences did not complement the virulence of a
strain defective for
gene activation. While editing the TalC EBE at the ATAC loci compromised TalC-mediated induction, multiplex edited lines with mutations at the
and ATAC loci remained essentially susceptible to African
strains. Overall, this work indicates that ATAC is a probable TalC off-target locus but nonetheless documents the first example of divergent transcription activation by a native TALE during infection.
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