Fig. 5.

PIF5 does not interact with UVR8 but binds to COP1, destabilising PIF5. a Co-IP assay showing that PIF5 does not interact with UVR8 in planta. LerPIF5Ox Seedlings were grown for 10 days in16 h light/8 h dark cycles before transfer at dawn to high R:FR ± UV-B for 1 h. Total protein extracts were immunoprecipitated with anti-HA beads and immunoblots analysed with anti-HA or anti-UVR8 antibodies. An anti-PHYB antibody was used as positive control for PIF5 immunoprecipitation. Ponceau stained Rubisco large subunit (rbcL) was used as a loading control. b Western blot of COP1 protein abundance in Ler and uvr8-1. Seedlings were grown as in (a) and immunoblots probed with anti-COP1 and anti-UGPase antibodies. c Co-IP assay showing that PIF5 interacts with COP1 in the presence and absence of UV-B. LerPIF5Ox and WT seedlings were grown as in (a). Immunoblots were probed with anti-COP1 or anti-HA antibodies. Ponceau staining of the Rubisco large subunit (rbcL) was used as a loading control. d Quantification of IP/input protein ratio in (c) Mean values from three biological repeats are shown. Bars represent SE. e Co-IP assay performed as in (c) showing increased PIF5-COP1 complex in uvr8-1 mutants in UV-B. LerPIF5Ox and uvr8-1PIF5Ox seedlings were grown as in (a). f Quantification of IP/input protein ratio in (e). Mean values from three biological repeats are shown. Bars represent s.e.m. g, h Western blots of PIF5 protein abundance in 10-day-old Col-0 and cop1-4 seedlings. Plants were grown in 16 h light/8 h dark cycles before transfer at dawn to high R:FR (WL) ± UV-B (e) or low R:FR (FR) ± UV-B (f) for 2 h. Immunoblots were probed with an anti-PIF5 antibody. Ponceau staining of the Rubisco large subunit (rbcL) was used as a loading control. Source data are provided as a source data file