Figure 6.

The interaction between CaNAC2c and CaNAC029 and its effect on transcription of immunity or thermotolerance related genes by CaNAC2c. A, BiFC confirmation of the interaction between CaNAC2c and CaNAC029 in N. benthamiana leaves infiltrated with Agrobacterium cells bearing CaNAC2c-YFPC+CaNAC029-YFPN or CaNAC2c-YFPN+CaNAC029-YFPC constructs, NbH3 (histone H3)-CFP was used to indicate the nucleus. Cyan fluorescence and yellow fluorescence, visible light, and merged images were taken on a confocal microscope at 48 hpi. Bars = 25 μm. B, Interaction between CaNAC2c and CaHSP70 in vivo, as determined by Co-IP assay. Proteins were isolated from pepper leaves transiently overexpressing CaNAC2c-Myc and CaNAC029-HA, and CaNAC2c-Flag and its interacting partners were immunoprecipitated with antibody of Flag, the presence of CaNAC029 in the protein complex was assayed by western blotting using antibody of HA. C, The effect of CaNAC029 silencing on the HR cell death triggered by transient overexpression of CaNAC2c under room temperature and upon HTS challenge. D, CaNAC029 cannot bind to the promoter of CaHSFA5 upon RSI or HTS, and co-transient overexpression of CaNAC029 and CaNAC2c inhibit the binding of CaNAC2c to CaHSFA5 promoters at room temperature. E, The effect of transient overexpression of CaNAC029 on the regulation of CaHSFA5 and CaDEF1 by CaNAC2c by RT-qPCR. F, The effect of CaNAC029 silencing on the regulation of CaHSFA5 and CaDEF1 by transient overexpression of CaNAC2c under room temperature and upon HTS challenge by RT-qPCR. In D, E, and F, data are shown as means ± standard error of four replicates. Different capital letters above the bars indicate significant differences between means (P < 0.01) by Fisher’s protected least-significant-difference (LSD) test.