MKP-1 modulates subcellulartranslocalization of AUF1. Rat macrophages were transduced with Ad.MKP-1 or control Ad.LacZ (300 MOI), or treated with HEPES buffered saline for 48 h. Primary bone marrow macrophages were either harvested from MKP-1 KO mice or WT mice. The macrophages were either unstimulated or stimulated with 1μg/ml of LPS (from Escherichia coli 0127:B8) for 2 or 4 h. The nuclear and cytoplasmic proteins were extracted and analyzed by Western blot assay. Cytoplasmic protein (30 μg) and nuclear protein (15 μg) was loaded on the gel. (a) MKP-1, p-JNK (p54, p46), p-p38, p-ERK (p44, p42), AUF1 (p45, p42, p40, p37), and TTP proteins expression in the cytoplasm vs. in the nucleus of rat macrophages. (b) AUF1 (p45, p42, p40, p37), and TTP proteins expression in the cytoplasm vs. in the nucleus of mouse bone marrow macrophages.The GAPDH (cytoplasmic protein) and Lamin A/C (nuclear protein) are loading controls for cytoplasmic protein and nuclear protein separately. (c) AUF1 and TTP protein density in rat macrophages transduced with AdMKP-1 (black bars), or Ad.LacZ (white bars) or treated with HEPES buffered saline (gray bars) (n=3, *p<0.05). (d) AUF1 and TTP protein density in bone marrow (BM) macrophages from MKP-1 KO mice (black bars) and WT mice (white bars)(n=3, *p<0.05). The AUF1 and TTP protein density/mm2 was analyzed by Quantity One software and normalized by cytoplasmic protein GAPDH or nuclear protein Lamin A/C respectively. These data are representative of three separate experiments.