Fig. 5: TRPM8 interacts directly with the N-terminal region of AR independently of the polyQ tract.

a GST pull-down assay of in vitro translated AR and GST, GST fused to the TRPM8 N-terminal tail (GST-Nt) or to C-terminal tail (GST-Ct), as indicated. For the input of the GST pull-down assay, 10% of the in vitro translated AR was used. Right: plot compares the AR-TRPM8 tail interactions normalized to the input. Cartoon (inset) illustrates the GST-Nt and GST-Ct purified fragments. b GST pull-down assay of in vitro translated AR and GST, GST fused to GST-Nt or to GST-Ct. In vitro translated AR was treated with 1, 10 or 100 nM testosterone during incubation with the N- or C-terminal tail of TRPM8. Right: plot compares the AR-TRPM8 tail interactions normalized to the input. N = 3 independent experiments. c Schematic representation of major AR domains: NTD, DBD and LBD. GST pull-down assay of in vitro translated AR-DBD/LBD tails d or AR N-terminal tail e with GST, GST-Nt or GST-Ct, as indicated. f GST pull-down assay of in vitro translated AR mutants harboring polyQ tract deletions (hARΔpolyQ, hARΔQ5, hARΔpolyQQ5, hARΔpolyQQ6 or hARΔpolyQQ5Q6, as indicated) with GST, GST-Nt or GST-Ct. Right: plot compares the interactions between different AR mutants and TRPM8 termini normalized to the input. For each pull-down, N = 3 independent experiments.**P < 0.01, ***P < 0.005 (ANOVA)