Abstract
WRKY proteins play important roles in transcriptional reprogramming in plants in response to various stresses including pathogen attack. In this study, we functionally characterized a rice WRKY gene, OsWRKY67, whose expression is upregulated against pathogen challenges. Activation of OsWRKY67 by T-DNA tagging significantly improved the resistance against two rice pathogens, Magnaporthe oryzae and Xanthomonas oryzae pv. oryzae (Xoo). Reactive oxygen species (ROS) rapidly accumulated in OsWRKY67 activation mutant lines in response to elicitor treatment, compared with the controls. Overexpression of OsWRKY67 in rice confirmed enhanced disease resistance, but led to a restriction of plant growth in transgenic lines with high levels of OsWRKY67 protein. OsWRKY67 RNAi lines significantly reduced resistance to M. oryzae and Xoo isolates tested, and abolished XA21-mediated resistance, implying the possibility of broad-spectrum resistance from OsWRKY67. Transcriptional activity and subcellular localization assays indicated that OsWRKY67 is present in the nucleus where it functions as a transcriptional activator. Quantitative PCR revealed that the pathogenesis-related genes, PR1a, PR1b, PR4, PR10a, and PR10b, are upregulated in OsWRKY67 overexpression lines. Therefore, these results suggest that OsWRKY67 positively regulates basal and XA21-mediated resistance, and is a promising candidate for genetic improvement of disease resistance in rice.
Highlights
The WRKY proteins possess a unique conserved domain, WRKYGQK, and can bind to the DNA sequence (T)(T)TGAC(C/T) called the W-box (Eulgem et al, 2000)
To elucidate OsWRKY67 function, two T-DNA activation tagging mutant plants with enhanced expression of OsWRKY67 were isolated from our T-DNA activation library and named OsW67-D1 and OsW67-D2 (Figure 1A, Top) (Jeon et al, 2000; Jeong et al, 2002)
We found that the OsW67-D1 and OsW67-D2 lines accumulated Reactive oxygen species (ROS) rapidly and highly in response to elicitor treatments (Figure 2)
Summary
The WRKY proteins possess a unique conserved domain, WRKYGQK, and can bind to the DNA sequence (T)(T)TGAC(C/T) called the W-box (Eulgem et al, 2000). A number of studies suggest that WRKYs play a key role in transcriptional reprogramming upon pathogen perception, as either an activator or a repressor (Pandey and Somssich, 2009; Rushton et al, 2010; Chen et al, 2012). WRKYs regulate expression of pathogenesis-related (PR) genes (Jimmy and Babu, 2015) and induce defense metabolism pathways (Jiang et al, 2017) directly or indirectly; they are a key family for molecular studies, as well as breeding programs. For instance, data accumulated from microarray analysis and RNA sequencing revealed that OsWRKYs dynamically respond to. Role of OsWRKY67 in Rice Disease Resistance various stimuli, including biotic and abiotic stresses (Ramamoorthy et al, 2008). A number of OsWRKY genes are induced by Magnaporthe oryzae and Xanthomonas oryzae pv. A number of OsWRKY genes are induced by Magnaporthe oryzae and Xanthomonas oryzae pv. oryzae (Xoo), the causal agents of rice blast and bacterial leaf blight disease, respectively (Ryu et al, 2006)
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