a, Western blotting analysis of etiolated 35S-IAA32-GFP line in either wild-type or tmk1, treated with auxin for indicated time points. Three biological repeats. b, Western blotting analysis of 35S-IAA28-MYC line treated with auxin for indicated time points; asterisk indicates non-specific band. Three biological repeats. c, The TIR1 inhibitor PEO-IAA did not affect auxin-mediated accumulation of IAA32 and IAA34. Three biological repeats. d, Level of IAA32 and IAA34 transcription was downregulated in tir1 afb2 afb3 mutant. Three biological repeats. Dots show data distribution. Two-sided t-test. Data are mean ± s.d. The expression level was standardized to 1 in wild-type seedlings. UBQ10 was used as internal control. e, The protein stability of IAA32 with or without co-expression of 35S–TMK1 C-terminus –HA. CHX treatment for indicated time periods. 35S–sGFP was used as a control. Three biological repeats. f, Western blot of proteins in protoplasts co-transfected with HBT–TMK1 C-terminus –HA and HBT–IAA14–GFP. Three repeats showed similar results. g, The protein stability of IAA32 with or without co-expression of 35S–TMK1 C-terminus –HA. CHX treatment for indicated time periods. 35S–sGFP was used as a control. Three biological repeats. h, Confocal microscopy of IAA32–GFP protein in the apical hook of a tmk1 mutant with gTMK1-FLAG or gTMK1(K616E)-FLAG. Scale bars, 50 μm. Three biological repeats. i, Representative images of apical hooks in Col-0, tmk1-1, gTMK1-FLAG;tmk1-1 and gTMK1(K616E)-FLAG;tmk1-1 at indicated time points after germination, Scale bars, 500 μm. Three independent repeats showed similar results. j, Quantification of apical-hook curvature at the time points corresponding to those shown in i. n = 15 biologically independent seedlings; data are mean ± s.e.m. For western blot images a–c and g, the same membrane was stripped and blotted with anti-actin antibodies as a loading control. For f, IAA14–GFP and TMK1 C-terminus –HA were run separately (with the same amount of protein per 10-μl sample) owing to the similar molecular mass of these two proteins. The same membrane of IAA14–GFP was stripped and blotted by anti-actin antibodies as a loading control. Source Data