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. 2005 Sep;25(18):8259–8272. doi: 10.1128/MCB.25.18.8259-8272.2005

FIG. 3.

FIG. 3.

NDR1 phosphorylated on Thr444 is mostly cytoplasmic. A. COS-7 cells expressing HA-tagged wild-type (wt) (top panels), Thr444Ala (T444A) (middle panels), or kinase-dead (K118A) (bottom panels) NDR1 were either left untreated (− OA) or incubated with okadaic acid (+ OA) before processing for immunofluorescence using anti-HA 12CA5 (red) and anti-T444-P (green) antibody. Nuclei are indicated in blue. All pictures were taken using the same settings, which had been adjusted to signals of cells overexpressing NDR1. B. COS-7 cells expressing HA-tagged NDR1 were subjected to biochemical cell fractionation after OA treatment (T, total; N, nuclear; C, cytoplasmic; M, membrane) and processed for immunoblotting with anti-T444-P (top panel), anti-HA 12CA5 (top middle panel), anti-lamin A/C (middle panel), anti-α-tubulin (bottom middle panel), and anti-p63/Climp (bottom panel) antibodies. C. After OA treatment, U2-OS cells were subjected to biochemical cell fractionation (T, total; N, nuclear; C, cytoplasmic; M, membrane) and processed for immunoblotting with anti-T444-P (top panel), anti-NDR1 (top middle panel), anti-lamin A/C (middle panel), anti-α-tubulin (bottom middle panel), and anti-p63/Climp (bottom panel) antibodies.