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. Author manuscript; available in PMC: 2009 Aug 22.
Published in final edited form as: Mol Cell. 2008 Aug 22;31(4):607–613. doi: 10.1016/j.molcel.2008.08.003

Figure 1. The COP1/SPA1 complex associates with an E3 activity for phyA ubiquitination.

Figure 1

(A) TAPSPA1 co-purified proteins from seedlings following the purification scheme (right) were resolved by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and silver-stained. Immunoblots with anti-myc (for TAPSPA1) and anti-COP1 antibodies are presented (middle). The box shows COP1 peptide fragments identified by mass spectrometry analysis. GFP, TAPGFP; SPA1, TAPSPA1; IgG, Immunoglobulin G; 3C–P, 3C protease. (B and C) phyA ubiquitination by TSC in vitro. Purified TSC (B) and an analytical scale TSC fraction (C) as well as their corresponding TAPGFP controls were assayed for E3 activity on GSTphyA. Ubiquitinated products were detected by immunoblotting (WB) with anti-phyA and ubiquitin (Ub) antibodies (B) or with anti-phyA and Flag antibodies (C). phyA was recovered by immunoprecipitation (IP) with anti-GST antibodies (C). The open and closed arrowheads indicate positions of apparently mono-ubiquitinated and unmodified phyA, respectively.