Akt-mediated GSK-3β inhibition prevents migration of polyamine-depleted intestinal epithelial cells via Rac1

The rapid migration of intestinal epithelial cells (IEC) is important for the healing of mucosal wounds. We have previously shown that polyamine depletion inhibits migration of IEC-6 cells. Akt activation and its downstream target GSK-3β have been implicated in the regulation of migration. Here we investigated the significance of elevated phosphatidylinositol 3-kinase (PI3K)/Akt signaling on migration of polyamine-depleted cells. Polyamine-depleted cells had high Akt (Ser473) and GSK-3β (Ser9) phosphorylation. Pretreatment with 20 μM LY294002 (PI3K inhibitor) for 30 min inhibited phosphorylation of Akt, increased migration by activating Rac1 in polyamine-depleted IEC-6 cells, and restored the actin structure similar to that in cells grown in control medium. Treatment of cells with a GSK-3β inhibitor (AR-A014418) altered the actin cytoskeleton and inhibited migration, mimicking the effects of polyamine depletion. Thus, our results indicate that sustained activation of Akt in response to polyamine depletion inhibits migration through GSK-3β and Rac1. Received 25 August 2006; received after revision 3 October 2006; accepted 16 October 2006     

PINK1 stimulates interleukin-1β-mediated inflammatory signaling via the positive regulation of TRAF6 and TAK1

Parkinson's disease (PD) is characterized by a progressive loss of dopaminergic neurons in the substantia nigra. The cause of neuronal death in PD is largely unknown, but several genetic loci, including PTEN-induced putative kinase 1 (PINK1), have been linked to early onset autosomal recessive forms of familial PD. PINK1 encodes a serine/threonine kinase, which phosphorylates several substrates and consequently leads to cell protection against apoptosis induced by various stresses. In addition, research has shown that inflammation largely contributes to the pathogenesis of PD, but the functional link between PINK1 and PD-linked neuroinflammation remains poorly understood. Therefore, in the present study, we investigated the functional role of PINK1 in interleukin (IL)-1β-mediated inflammatory signaling. We show that PINK1 specifically binds to TRAF6 and TAK1, and facilitates the autodimerization and autoubiquitination of TRAF6. PINK1 also enhances the association between TRAF6 and TAK1, phosphorylates TAK1, and stimulates polyubiquitination of TAK1. Furthermore, PINK1 leads to the potentiation of IL-1β-mediated NF-κB activity and cytokine production. These findings suggest that PINK1 positively regulates two key molecules, TRAF6 and TAK1, in the IL-1β-mediated signaling pathway, consequently up-regulating their downstream inflammatory events.     

Efficient cell proliferation and predominant accumulation of ɛ-globin mRNA in human leukemic K562 cells which produce mostly Hb Gower 1

Summary Long-term cultures of K562(S) cells in 50–75 M hemin allow the selection of hemin-resistant K562 cells together with cells which proliferate efficiently while fully induced to express the human embryonic globin genes, as the hemoglobin Gower 1 (₂₂) is the predominant hemoglobin produced. Our experiments demonstrate that these K562 cells accumulate mostly -globin mRNA (-globin mRNA/-globin mRNA=2.9) suggesting that the control of hemoglobin expression is at a pretranslational level.We thank Dr Irene Bozzoni (Centro degli Acidi Nucleici, Università di Roma) for the pXCR7 probe. Address for reprint request: R.G. Centro Studi Biochimici sul Morbo di Cooley, Via Borsari 46, I-44100 Ferrara.     

Overexpression of HAb18G/CD147 promotes invasion and metastasis via α3β1 integrin mediated FAK-paxillin and FAK-PI3K-Ca2+ pathways

Mechanism of HAb18G/CD147 underlying the metastasis process of human hepatoma cells has not been determined. In the present study, we found that integrin α3β1 colocalizes with HAb18G/CD147 in human 7721 hepatoma cells. The enhancing effect of HAb18G/CD147 on adhesion, invasion capacities and matrix metalloproteinases (MMPs) secretion was decreased by integrin α3β1 antibodies (p<0.01). The expressions of integrin downstream molecules including focal adhesion kinase (FAK), phospho-FAK (p-FAK), paxillin, and phospho-paxillin (p-paxillin) were increased in human hepatoma cells overexpressing HAb18G/CD147. Deletion of HAb18G/CD147 reduces the quantity of focal adhesions and rearranges cytoskeleton. Wortmannin and LY294002, specific phosphatidylinositol kinase (PI3K) inhibitors, reversed the effect of HAb18G/CD147 on the regulation of intracellular Ca²⁺ mobilization, significantly reducing cell adhesion, invasion and MMPs secretion potential (p<0.01). Together, these results suggest that HAb18G/CD147 enhances the invasion and metastatic potentials of human hepatoma cells via integrin α3β1-mediated FAK-paxillin and FAKPI3K-Ca²⁺ signal pathways. Received 5 June 2008; received after revision 16 July 2008; accepted 23 July 2008