Protein kinase D1 mediates class IIa histone deacetylase phosphorylation and nuclear extrusion in intestinal epithelial cells: role in mitogenic signaling.

We examined whether class IIa histone deacetylases (HDACs) play a role in mitogenic signaling mediated by protein kinase D1 (PKD1) in IEC-18 intestinal epithelial cells. Our results show that class IIa HDAC4, HDAC5, and HDAC7 are prominently expressed in these cells. Stimulation with ANG II, a potent mitogen for IEC-18 cells, induced a striking increase in phosphorylation of HDAC4 at Ser(246) and Ser(632), HDAC5 at Ser(259) and Ser(498), and HDAC7 at Ser(155). Treatment with the PKD family inhibitors kb NB 142-70 and CRT0066101 or small interfering RNA-mediated knockdown of PKD1 prevented ANG II-induced phosphorylation of HDAC4, HDAC5, and HDAC7. A variety of PKD1 activators in IEC-18 cells, including vasopressin, lysophosphatidic acid, and phorbol esters, also induced HDAC4, HDAC5, and HDAC7 phosphorylation. Using endogenously and ectopically expressed HDAC5, we show that PKD1-mediated phosphorylation of HDAC5 induces its nuclear extrusion into the cytoplasm. In contrast, HDAC5 with Ser(259) and Ser(498) mutated to Ala was localized to the nucleus in unstimulated and stimulated cells. Treatment of IEC-18 cells with specific inhibitors of class IIa HDACs, including MC1568 and TMP269, prevented cell cycle progression, DNA synthesis, and proliferation induced in response to G protein-coupled receptor/PKD1 activation. The PKD1-class IIa HDAC axis also functions in intestinal epithelial cells in vivo, since an increase in phosphorylation of HDAC4/5 and HDAC7 was demonstrated in lysates of crypt cells from PKD1 transgenic mice compared with matched nontransgenic littermates. Collectively, our results reveal a PKD1-class IIa HDAC axis in intestinal epithelial cells leading to mitogenic signaling.