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The high mobility group box 1 (HMGB1) protein, a non-histone nuclear factor, is overexpressed and localizes to the cytoplasm in some cancer cells. However, the mechanism of cytoplasmic HMGB1 transport, extracellular secretion, and its role in cancer progression is not clear. Here we show that phosphorylated HMGB1 localizes in the cytoplasm of colon cancer cells and that protein kinase C (PKC) signaling is directly involved in the translocation of HMGB1 from the nucleus to the cytoplasm. In order to simulate the phosphorylation state of activated HMGB1, we mutated serine residues of nuclear localization signals (NLS) to glutamic acid. Concurrent mutations at six serine residues (35, 39, 42, 46, 53, and 181) induced nuclear to cytoplasmic transport of the mutant HMGB1, which was detected in the culture medium. Among these six serine residues, mutations at 35, 39 and 42, the predicted phosphorylation target sites by PKC, were critical for nuclear to cytoplasmic transport. Inactivation of PKC inhibited the nuclear to cytoplasmic transport of HMGB1. We also observed that secretion of HMGB1 correlated with activation of the matrix metalloproteinase 2 (MMP2) pathway and increased cancer cell invasiveness. Our results suggest that HMGB1 is transported to the cytoplasm due to PKC signaling-mediated phosphorylation, is subsequently secreted from the cell, and plays a role in tumor progression through the activation of genes related to cell migration.

99th AACR Annual Meeting-- Apr 12-16, 2008; San Diego, CA