Roles of WRKY Transcription Factors in Response to Sri Lankan Cassava Mosaic Virus Infection in Susceptible and Tolerant Cassava Cultivars

Abstract

Cassava mosaic disease (CMD) is caused by viruses such as Sri Lankan cassava mosaic virus (SLCMV). It poses a significant threat to the cassava (Manihot esculenta) yield in Southeast Asia. Here, we investigated the expression of WRKY transcription factors (TFs) in SLCMV-infected cassava cultivars KU 50 (tolerant) and R 11 (susceptible) at 21, 32, and 67 days post-inoculation (dpi), representing the early, middle/recovery, and late infection stages, respectively. The 34 identified WRKYs were classified into the following six groups based on the functions of their homologs in the model plant Arabidopsis thaliana (AtWRKYs): plant defense; plant development; hormone signaling (abscisic, salicylic, and jasmonic acid); reactive oxygen species production; basal immune mechanisms; and other related hormones, metabolites, and abiotic stress responses. Regarding the protein interactions of the identified WRKYs, based on the interactions of their homologs (AtWRKYs), WRKYs increased reactive oxygen species production, leading to salicylic acid accumulation and systemic acquired resistance (SAR) against SLCMV. Additionally, some WRKYs were involved in defense-related mitogen-activated protein kinase signaling and abiotic stress responses. Furthermore, crosstalk among WRKYs reflected the robustly restricted viral multiplication in the tolerant cultivar, contributing to CMD recovery. This study highlights the crucial roles of WRKYs in transcriptional reprogramming, innate immunity, and responses to geminivirus infections in cassava, providing valuable insights to enhance disease resistance in cassava and, potentially, other crops.

Keywords: Sri Lankan cassava mosaic virus; WRKYs; phenotypic variations; plant defense mechanisms.

Figure 1

Venn diagrams of all identified WRKYs in this study, which involved 34 WRKYs identified at 21, 32, and 67 dpi in SLCMV-infected (a) KU 50 and (b) R 11.

Figure 2

Heatmap of identified WRKYs by cultivar and SLCMV infection stage based on Fisher’s least significant difference tests (p < 0.05).

Figure 3

Phylogenetic tree of Arabidopsis thaliana WRKYs (AtWRKYs) and the identified WRKYs (with NCBI accession numbers) based on amino acid sequence alignment. The tree was constructed using the NGPhylogeny platform and adjusted using the Interactive Tree of Life (ITOL) tool (bootstrap values were superimposed on branches with <50% support). Classification into three groups (groups 1, 2 and 3), with five subgroups for group 2 (2a–e), was primarily based on the zinc finger motif [21]. Red boxes represent nine key WRKYs that were selected based on the DEG analysis results.

Figure 4

Venn diagrams of (a) upregulated and (b) downregulated differentially expressed genes (DEGs; p < 0.01 and log₂(fold change) ≤ 1.0) at 32 and 67 dpi in SLCMV-infected KU 50 and R 11.

Figure 5

Protein–protein interaction network of WRKYs based on STITCH v5 analysis. Red circles highlight the WRKYs identified in this study.

Figure 6

Summary of functions and interacting proteins of nine selected WRKY DEGs at 32 and 67 dpi in SLCMV-infected KU 50 (tolerant) and R 11 (susceptible). Blue, orange, and green boxes represent WRKYs involved in basal immunity (such as PTI and ETI pathways), abiotic stress responses, and other hormones, respectively. Regarding the blue boxes, the seven WRKYs (WRKY22, -24, -40, -43, -47, -57, and -75) participate in plant basal immunity responses, interacting with the ETI or PTI signaling pathways. Arrows represent the interacting proteins/corresponding genes and the plant responses. These WRKYs are up- and downregulated in response to SLCMV infection. Regarding the orange and green boxes, WRKY23 and -39 regulate abiotic stress responses and hormone biosynthesis, respectively.