Fig. 3: S744 is required for 14-3-3 binding and early signalling events. | Nature Communications

Fig. 3: S744 is required for 14-3-3 binding and early signalling events.

From: Regulation of plant phototropic growth by NPH3/RPT2-like substrate phosphorylation and 14-3-3 binding

Fig. 3

a Confocal images of hypocotyl cells of etiolated nph3 seedlings expressing GFP-NPH3 (NPH3) or phosphorylation site mutants S744A S746A, S744D S746D, S744A and S746A. Seedlings were scanned immediately (scan 1) and again after repeat scanning with the 488 nm laser (scan 5). GFP is shown in green and autofluorescence in magenta. Bar, 50 µm. b Immunoblot analysis of total protein extracts from etiolated nph3 seedlings expressing GFP-NPH3 (NPH3) or phosphorylation site mutants maintained in darkness (D) or irradiated with 20 µmol m−2 s−1 blue light for 15 min (L). Protein extracts were probed with anti-NPH3 antibodies. c Far-western blot analysis of anti-GFP immunoprecipitations from etiolated nph3 seedlings expressing GFP-NPH3 (NPH3) or phosphorylation site mutants (S744A or S746A) maintained in darkness (D) or irradiated with 20 µmol m−2 s−1 of blue light for 15 min blue light (L). GST-tagged 14-3-3 isoform Epsilon was used as the probe. Blots were probed with an anti-NPH3 antibody as a loading control (bottom panel). Experiments were repeated at least twice with similar results.

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