Rice WRKY13 TF protein binds to motifs in the promoter region to regulate downstream disease resistance-related genes.

In plants, pathogen resistance is brought about by the binding of certain transcription factor (TF) proteins to the cis-elements of certain target genes. These cis-elements are present upstream in the motif of the promoters of each gene. This ensures the binding of a specific TF to a specific promoter, therefore regulating the expression of that gene. Therefore, the study of each promoter sequence of all the rice genes would help identify the target genes of a specific TF. Rice 1 kb upstream promoter sequences of 55,986 annotated genes were analyzed using the Perl program algorithm to detect WRKY13 binding motifs (bm). The resulting genes were grouped using Gene Ontology and gene set enrichment analysis. A gene with more than 4 TF bm in their promoter was selected. Ten genes reported to have a role in rice disease resistance were selected for further analysis. Cis-acting regulatory element analysis was carried out to find the cis-elements and confirm the presence of the corresponding motifs in the promoter sequences of these genes. The 3D structure of WRKY13 TF and the corresponding ten genes were built, and the interacting residues were determined. The binding capacity of WRKY13 to the promoter of these selected genes was analyzed using docking studies. WRKY13 was considered for docking analysis based on the prior reports of autoregulation. Molecular dynamic simulations provided more details regarding the interactions. Expression data revealed the expression of the genes that helped provide the mechanism of interaction. Further co-expression network helped to characterize the interaction of these selected disease resistance-related genes with the WRKY13 TF protein. This study suggests downstream target genes that are regulated by the WRKY13 TF. The molecular mechanism involving the gene network regulated by WRKY13 TF in disease resistance against rice fungal pathogens is explored.

Proteome Responses to Individual Pathogens and Abiotic Conditions in Rice Seedlings.

Rice plants under field conditions experience various biotic and abiotic stresses and are adapted to survive using a molecular cross-talk of genes and their protein products based on the severity of a given stress. Seedlings of cultivated variety ASD16 (resistant to fungal disease, blast; tolerant to abiotic stress, salinity) were subjected to salt, drought, high temperature and low temperature stress as well as infection by Rhizoctonia solani and Xanthomonas oryzae pv. oryzae (causing reemerging diseases such as sheath blight and leaf blight), respectively, the sheath blight and bacterial leaf blight pathogens. Leaf proteome was analyzed using two-dimensional electrophoresis and differentially expressed proteins were identified using mass spectrometry. In addition to many other differentially expressed proteins, acidic endochitinase was found to be upregulated during fungal infection and drought treatment, and a germin-like protein upregulated during fungal infection and high temperature stress. These two proteins were further validated at the gene expression level using reverse transcription-PCR in dual stress experiments. Pot culture plants were subjected to fungal infection followed by drought and drought followed by fungal infection to validate chitinase gene expression. Similarly, plants subjected to fungal infections followed by high temperature stress and vice versa were used to validate the expression of germin-like protein-coding gene. The results of the present study indicate that chitinase and germin-like protein are potential targets for further exploration to develop rice plants resistant or tolerant to biotic and abiotic stresses.