No abstract

Mesenchymal stem cell (MSC)-based transplantation is a promising therapeutic approach for bone regeneration and repair. In the realm of therapeutic bone regeneration, the defect or injured tissues are frequently inflamed with an abnormal expression of inflammatory mediators. Growing evidence suggests that proinflammatory cytokines inhibit osteogenic differentiation and bone formation. Thus, for successful MSC-mediated repair, it is important to overcome the inflammation-mediated inhibition of tissue regeneration. In this study, using genetic and chemical approaches, we found that proinflammatory cytokines TNF and IL-17 stimulated IκB kinase (IKK)–NF-κB and impaired osteogenic differentiation of MSCs. In contrast, the inhibition of IKK–NF-κB significantly enhanced MSC-mediated bone formation. Mechanistically, we found that IKK–NF-κB activation promoted β-catenin ubiquitination and degradation through induction of Smurf1 and Smurf2. To translate our basic findings to potential clinic applications, we showed that the IKK small molecule inhibitor, IKKVI, enhanced osteogenic differentiation of MSCs. More importantly, the delivery of IKKVI promoted MSC-mediated craniofacial bone regeneration and repair in vivo. Considering the well established role of NF-κB in inflammation and infection, our results suggest that targeting IKK–NF-κB may have dual benefits in enhancing bone regeneration and repair and inhibiting inflammation, and this concept may also have applicability in many other tissue regeneration situations.

SUMMARY Abnormal activation of Wnt/β-catenin-mediated transcription is associated with a variety of human cancers. Here we report that LATS2 inhibited oncogenic Wnt/β-catenin-mediated transcription by disrupting the β-catenin/BCL9 interaction. LATS2 directly interacted with β-catenin and to be present on Wnt target gene promoters. Mechanistically, LATS2 inhibited the interaction between BCL9 and β-catenin and subsequent recruitment of BCL9, independent of LATS2 kinase activity. LATS2 was down-regulated and inversely correlated with the levels of Wnt target genes in human colorectal cancers. Moreover, nocodazole, an anti-microtubule drug, potently induced LATS2 to suppress tumor growth in vivo by targeting β-catenin/BCL9. Our results suggest that LATS2 is not only a key tumor suppressor in human cancer, but may also be an important target for anti-cancer therapy.

NF-B, a pleiotropic transcription factor, activates a variety of gene transcription factors, including cytokines, angiogenesis modifiers, cell adhesion molecules, and antiapoptotic factors (29,34,(45)(46)(47)(48). The classical form of NF-B is a heterodimer consisting of p65/RelA and p50 subunits. In most unstimulated normal cells, NF-B is retained in the cytoplasm by IB inhibitory proteins. The stimulation of cells with tumor necrosis factor alpha (TNF-␣), interleukin-1, or various bacterial products activates the IB kinase (IKK) complex, leading to the phosphorylation of IB family proteins. Phosphorylated IB is polyubiquitinated by ubiquitin ligase and subsequently degraded by 26S proteasome machinery, resulting in translocation of NF-B to the nucleus to induce gene expression (31,32,(45)(46)(47)(48). Growing evidence suggests that aberrant activation of NF-B is a common feature of cancer cells (37,38). NF-B has been shown to promote tumor invasion and metastasis by regulating the expression of chemokines, cellular adhesion molecules, and matrix metalloproteinases (2,9,30,31,33). Previously we demonstrated that constitutive activation of NF-B promotes osteolytic breast cancer bone metastasis (33). Improved understanding of the molecular mechanisms that control NF-B-mediated gene regulation may lead to development of novel strategies to target NF-B signaling for inhibition of cancer progression and metastasis.When NF-B moves into the nucleus, it interacts with the transcription coactivator CBP/p300 to activate transcription.While many studies have shown that posttranscriptional modification of p65 affects how it interacts with coactivators and corepressors, it is unknown how p65 is recruited to the NF-B target gene promoter to activate gene transcription. In general, transcriptional activation by liganded nuclear receptors and other regulated transcription factors requires the dismissal of a corepressor and subsequent recruitment of coactivators with intrinsic enzyme activities (36). Transducin ␤-like protein 1 (TBL1) and TBL1-related protein (TBLR1), two highly related F-box and WD-40 domain-containing proteins, are the intrinsic components of the NCoR corepressor complex (11,26,42,49,51). TBL1 and TBLR1 have been found to be important in ligand-induced activation by serving as specific adaptors for the recruitment of the ubiquitin-conjugating/19S proteasome complex, which mediates the exchange of corepressors for coactivators (35). TBL1 is also involved in p53-mediated degradation of ␤-catenin induced by DNA-damaging agents (28). Recently, TBL1 was reported to interact with CtBP and mediate its ubiquitylation and degradation, thus leading to a dismissal of corepressor complexes during gene activation (36).Aside from serving a crucial role in regulated gene expression, our recent findings revealed unique functions of TBL1 in canonical Wnt signaling. We found that TBL1 plays an important role in the recruitment of ␤-catenin to the Wnt target gene promoter to activate transcription (25). Interestingly, TBL1 has ...