Figure 1. PIN2 ubiquitylation upon gravistimulation (A) western blots performed with PIN2 immunoprecipitates derived from 6-d-old Col-0 roots grown on horizontally positioned roots that were probed with either anti-PIN2 (top) or anti-ubiquitin (bottom) antibodies. Samples were gravistimulated by turning plates clockwise at an angle of 90 degrees. A first sample was taken at time point 0' immediately before gravistimulation. The duration of gravistimulation is indicated below the blots. All details on protein extraction as well as reagents and conditions used, were according to Leitner et al.³ Two biological repetitions have been performed. Ig-specific signals are indicated by an asterisk. (B) Quantification of signal intensities performed with ImageJ software. Signals on scanned blots (A) were quantified and ubiquitin-specific signals were normalized to PIN2 signals (signals corresponding to PIN2 and ubiquitin-specific signals ranging from 75 kDa to 230 kDa were considered for quantification; Ig-specific bands visible on the ubiquitin blot were omitted). (C) Control immunoprecipitation performed as described in (A), but without gravistimulation. Samples were extracted from roots of vertically positioned Col-0 seedlings at time points indicated on top of the blots. Ig-specific signals are indicated by an asterisk.