Extended Data Fig. 3: In planta expression of SOX converts Pip to P6C.
(a) Confocal micrographs showing transient expression of ALD1, FMO1, SOX, or co-expression of ALD1 + FMO1 or ALD1 + SOX in Nicotiana benthamiana plants. (b-d) Levels of Pip (b), NHP (c) and P6C (d) in N. benthamiana plants transiently expressing ALD1, SOX, FMO1, ALD1 + SOX or ALD1 + FMO1. Plants transiently expressing empty vector were used as controls. (e-f) UHPLC-MS analysis showing extracted ion-chromatograms e) and corresponding mass spectra (f) of P2C and P6C. Tissue extracts were prepared from mock- and Pst-avrRpt2 inoculated Col-0 plants sampled 24 h post inoculation. P6C standard (std) used in this analysis was synthesized using an in vitro SOX-catalyzed enzymatic reaction. The samples were run on a Kinetex F5, 1.7 µm (2.1 mm x 150 mm) column. The error bars represent SD (n = 3 or 4 biological replicates). Asterisks denote a significant difference (multiple unpaired t test, **P < 0.002; ***P < 0.0002; ****P < 0.0001). The experiments described here were repeated at least twice with similar results.
