Figure 3.

PhSLFs selectively interact with PhS-RNases. (A) Yeast two-hybrid assays between PhSLFs and PhS-RNases. Cells of yeast strain AH109 containing various combinations of bait (BD fusions) and prey (AD fusions) were tested for their growth on selective medium SD/-Ade-His-Leu-Trp. The empty vectors pGBKT7 and pGADT7 were negative controls. (B) The β-galactosidase activity assay was used to further test the interaction of PhSLFs and PhS-RNases. The combination of empty pGBKT7 and pGADT7 was used as a negative control. (C) Co-immunoprecipitation assays between PhS_3L-SLF1::FLAG and PhS₃-RNase or PhS_3L-RNase using in vitro pollen germination system. Untreated pollen tubes were used as negative control (CK). Samples from SPI (treated with S₃S₃ SE), CPC (treated with S_3LS_3LSE) and CK pollen tubes were immunoprecipitated by FLAG antibody and detected by PhS-RNase antibody and FLAG antibody, respectively. Challenged but not immmunoprecipitated SPI, CPC, and CK pollen tube samples were loaded as input. cFBP was detected as input loading control. PT, pollen tube; SE, style extract.