Extended Data Figure 5: Ectopic expression of T474D and LIMYB confers tolerance to begomovirus infection.

a, Delayed onset of infection in the T474D-4- and T474D-6-overexpressing lines. Ecotype Col-0 plants, as well as the T474D-4- and T474D-6-overexpressing lines, and the NIK1-8 and G473V/T474A-overexpressing lines were infected with CaLCuV DNA by the biolistic method. The progression of the infection was monitored by PCR detection of viral DNA in the systemic leaves of the inoculated plants. The values represent the percentages of systemically infected plants at different dpi. Error bars, 95% confidence intervals (n = 3) based on bootstrap resampling replicates of four independent experiments. b, Upon symptom development, the T474D-induced transcriptome diverges from the infected Col-0 transcriptome. The mock-inoculated T474D-overexpressing lines exhibited a constitutively infected wild-type transcriptome at 10 dpi (Fig. 1a). Nevertheless, these T474D transgenic lines did not phenocopy the infected wild-type plants because they did not develop symptoms of viral infection. In fact, the wild-type plants displayed typical symptoms of CaLCuV infection at 21 dpi, such as leaf distortion, stunting with epinasty and chlorosis (Fig. 1e). The symptoms in the T474D-expressing lines at 21 dpi, however, were greatly attenuated, with no visible leaf distortion or chlorosis. To examine these phenotypes, we performed a Ward hierarchical clustering of the gene expression data (normalized by the trimmed mean of M-values (TMM) normalization method) from the Arabidopsis infection experiments at 21 dpi. The TMM normalization method assumes that the majority of genes are not differentially expressed, and it adjusts genes with larger read counts and lower variance on the logarithmic scale. The dendrogram provides two types of P values: AU (black) and BP (grey). The AU P value comes from multiscale bootstrap resampling, while the BP value represents normal bootstrap resampling. These P values were calculated by multiscale bootstrap resampling using the R-cran package pvclust with a cut-off of 0.05. These P values show the significance of the proximity of each gene expression experiment profile. The cluster analysis at 21 dpi indicated that when symptoms had developed in the infected Col-0 leaves, the T474D-induced transcriptome diverged from the infected Col-0 transcriptome. The mock T474D transcriptome formed a unique clade, while the infected T474D transcriptome was more closely related to the mock-inoculated Col-0 transcriptome. c, Reduced viral transcript accumulation in T474D-overexpressing lines at 21 dpi. RNA-sequencing data of viral gene transcripts in the systemic leaves of infected wild-type and T474D-overexpressing plants at 10 dpi and 21 dpi. d, The onset of infection is delayed in LIMYB-overexpressing lines. Ecotype Col-0 plants and LIMYB-overexpressing and limyb mutant lines were infected with CaLCuV DNA using the biolistic method. The progression of the infection was monitored by the PCR detection of viral DNA in the systemic leaves of the inoculated plants. The values represent the percentages of systemically infected plants at different dpi. Error bars, 95% confidence intervals (n = 3) based on bootstrap resampling replicates of four independent experiments.