Fig. 6. Lysine239 and/or lysine240 in the cytoplasmic domain of proAR are essential for targeting to the nuclear envelope. (A) Amino acid sequence of the cytoplasmic domain of proAR. (B) Schematic representation of mutations in K239 and K240 in the cytoplasmic domain (AR-m2). AR-m2 p indicates AR-m2 precursor protein. HeLa cells transfected with AR-m2 were incubated with or without TPA and immunostained with anti-AR-N pAb (red) and anti-TGN46 pAb (green). (C) Schematic representation of V5-tag inserted AR and its deletion mutants. AR-V5-C p, AR-V5-C10 p and AR-V5-C11 p indicate precursor proteins. (D,E) HeLa cells transiently expressing AR-V5-C, AR-V5-C10 or AR-V5-C11 were incubated with or without TPA, then immunostained with anti-V5 mAb and anti-AR-N pAb (D) or with anti-AR-N pAb and anti-lamin A/C mAb (E). Arrowheads indicate AR-expressing cells. (F) Cell lysates of HeLa cells expressing AR-V-C5, AR-V5-C10 or AR-V5-C11 were immunoprecipitated with anti-lamin A/C mAb. Cell lysates (upper) and precipitated proteins (lower) were analyzed with anti-V5 mAb. Scale bars: 5 µm.