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Fig. 3. ProAR interacts with A-type lamin. (A) HeLa cells transiently expressing wild-type AREG were incubated with or without TPA. Cells were stained with anti-AR-C pAb and anti-lamin A/C mAb. Arrowheads indicate AREG-expressing cells. Scale bars: 5 µm. (B) Lysates of HeLa cells transiently transfected with or without wild-type AREG and treated with TPA for 60 minutes, were subjected to immunoblotting using anti-lamin A/C mAb (left) or anti-lamin B pAb (right). The left lanes show standard proteins. (C) HeLa cells transiently expressing wild-type AREG were incubated with or without TPA for 60 minutes. The cell lysates were subjected to immunoblotting with an anti-AR-C pAb (lane 1 and lane 2). The cell lysates were immunoprecipitated with an anti-lamin A/C mAb or normal mouse IgG, followed by western blotting using anti-AR-C pAb and anti-lamin A/C mAb. Arrows indicate the molecular mass of lamin A/C-interacted (upper) and AR-CTF (lower). (D) Schematic representation of GST-fused lamin A deletion mutants. (E) The cell lysates from cells transiently expressing wild-type AREG were incubated with GST or GST-lamin A derivatives, which were pulled down by glutathione-Sepharose beads. Bound proteins were analyzed by western blotting using the anti-AR-C pAb (upper panel). GST fusion proteins were analyzed by SDS-PAGE and stained with CBB (lower panel).
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