Extended Data Fig. 7: Analysis of LURE1 nitrosation.
From: FERONIA controls pectin- and nitric oxide-mediated male–female interaction
a, Coomassie-blue-stained gel of typical MBP and MBP–LURE1 preparations used for nitrosation assays. b, Ponceau-S-stained protein blot for detection of TMT-labelled MBP–LURE1 (arrow); immunoblot is shown in Fig. 4e. c, Ponceau-S-stained protein blot for detection of TMT-labelled MBP–full-length LURE1 (arrow); immunoblot is shown in Fig. 4e. d, Coomassie blue-stained purified MBP–full-length LURE1 after TMT-labelling reaction (arrow). The recombinant protein with LURE1 signal peptide tended to break down during the labelling procedure, producing a lower band at the MBP molecular weight range. The MBP-full LURE1 protein bands indicated by the arrow were excised for mass spectrometry analysis shown in Fig. 4e, g, h. e, LC–MS/MS spectrum showing TMT–Cys58-containing peptides, and LURE1.2 amino acid sequence highlighting Cys58. f, LURE1p::LURE1–GFP localization in ovules from unpollinated (BP) or pollinated (AP) pistils for comparison with LURE1(C17A)–GFP localization (shown in Fig. 4i). As shown in Fig. 4a, LURE1–GFP typically located in the filiform apparatus in ovules from unpollinated pistils, and delocalized to the cytoplasm of synergid cells in ovules from pollinated pistils. Box plots: centre line, median; box limits, lower and upper quartiles; dots, individual data points; whiskers, highest and lowest data points.
