NF-YA transcription factors suppress jasmonic acid-mediated antiviral defense and facilitate viral infection in rice
Fig 5
OsNF-YAs interact with OsMYC2/3.
A. Interaction of OsNF-YAs and OsMYC2/3 proteins in a yeast two-hybrid assay. The CDS for OsNF-YAs and OsMYCs were introduced into pGBKT7 and pGADT7 vectors, respectively. Yeast cells co-expressing OsNF-YAs and OsMYCs were grown on selective media SD/-Trp/-Leu (SD-L-T), and interactions were tested with SD/-Trp/-Leu/-His/-Ade (SD-L-T-H-A). Pictures were taken after 3 days incubation at 30°C. B. Co-IP assays confirming that OsNF-YA1 interacts with OsMYC2. OsMYC2-MYC was co-expressed with OsNF-YA1-FLAG or GFP-FLAG in N. benthamiana leaves. Total proteins were extracted and immunoprecipitated by anti-FLAG magnetic beads, and then detected with anti-MYC antibody. C. Co-IP assays confirming that OsNF-YA2 interacts with OsMYC2. OsMYC2-MYC was co-expressed with OsNF-YA2-GFP or GFP-FLAG in N. benthamiana leaves. Total proteins were extracted and immunoprecipitated by anti-GFP magnetic beads then detected with anti-MYC antibody. D. Co-IP assays confirming that OsNF-YA1 interacts with OsMYC3. OsMYC3-MYC was co-expressed with OsNF-YA1-FLAG or GFP-FLAG in N. benthamiana leaves. Total proteins were extracted and immunoprecipitated by anti-FLAG magnetic beads then detected with anti-MYC antibody. E. Co-IP assays confirming that OsNF-YA2 interacts with OsMYC3. OsMYC3-FLAG was co-expressed with OsNF-YA2-MYC or GFP-FLAG in N. benthamiana leaves. Total proteins were extracted and immunoprecipitated by anti-FLAG magnetic beads then detected with anti-MYC antibody. F-I. Results from LCI assays, and the measurements of luciferase activity due to LUC reconstitution for the different combinations, showing that OsNF-YA1 and OsNF-YA2 interact with OsMYC2 and OsMYC3 proteins in N. benthamiana leaves. J. BiFC assay showing interactions between OsNF-YA1/OsNF-YA2 and OsMYC2/3 in N. benthamiana cells. OsNF-YA1/OsNF-YA2 and OsMYC2/3 were fused with the N-terminal of YFP (nYFP) and the C-terminal of YFP (cYFP), respectively. Scale bar, 50 μm.