Biomedical subjects

Cun-Yu Wang

Publications and source records attributed to Cun-Yu Wang.

30 records · Page 2Linked to original sources

PubMed2003

Roles for homotypic interactions and transautophosphorylation in IkappaB kinase beta IKKbeta) activation [corrected].

The nuclear factor kappaB (NF-kappaB)/Rel family of transcription factors participates in a wide range of biological activities including inflammation, immunity, and apoptosis. NF-kappaB is kept inactive in the cytoplasm in unstimulated cells by virtue of the masking of its nuclear localization sequence by bound IkappaB protein. Cellular stimuli trigger the destruction of IkappaB proteins and the liberation of NF-kappaB to enter the nucleus and activate gene expression. A multisubunit IkappaB kinase complex (IKK) phosphorylates IkappaB proteins and mediates the activation of NF-kappaB by proinflammatory stimuli such as tumor necrosis factor alpha. Phosphorylation of IkappaB proteins triggers their polyubiquitination and their subsequent recognition and degradation by the proteasome. The IKK complex contains two catalytic subunits, IKKalpha and IKKbeta, and a noncatalytic subunit, NF-kappaB essential modifier/IKKgamma. IKK activation depends upon the phosphorylation of residues in the activation loop of IKKbeta and the subsequent activation of IKKbeta kinase activity. However, the events contributing to IKKbeta phosphorylation are not well understood. Here, we present evidence that the activation of IKKbeta depends on its ability to form homotypic interactions and to transautophosphorylate. We find that an intact leucine zipper in IKKbeta is necessary for homotypic interactions, kinase activation, and phosphorylation on its activation loop. Enforced oligomerization of an IKKbeta mutant defective in forming homotypic interactions restores kinase activation. Homotypic interactions allow IKKbeta molecules to transautophosphorylate one another on their activation loops. Finally, the oligomerization of IKKbeta is stimulated by tumor necrosis factor alpha in cultured cells. Our findings support a model whereby ligand-induced homotypic interactions between IKKbeta molecules result in IKKbeta phosphorylation and consequently IKK activation.

Cell Line↗

PubMed2003

A role for NF-kappaB essential modifier/IkappaB kinase-gamma (NEMO/IKKgamma) ubiquitination in the activation of the IkappaB kinase complex by tumor necrosis factor-alpha.

NEMO (NF-kappaB essential modifier)/IKKgamma (IkappaB kinase-gamma) is required for the activation of the IkappaB kinase complex (IKK) by inflammatory stimuli such as tumor necrosis factor (TNF-alpha). Here we show that TNF-alpha stimulates the ubiquitination of NEMO in a manner that does not appear to target it for degradation and that is impaired by mutations in the NEMO zinc finger. Mutations of the zinc finger are found in patients with hypohidrotic ectodermal dysplasia with immunodeficiency (HED-ID) and lead to the impairment of TNF-alpha-stimulated IKK phosphorylation and activation. In addition, the ubiquitination of NEMO is mediated by c-IAP1, an inhibitor of apoptosis protein that is a component of the TNF receptor signaling complex. Thus, the ubiquitination of NEMO mediated by c-IAP1 likely plays an important role in the activation of IKK by TNF-alpha. Also, defective NEMO ubiquitination may be responsible for the impaired cellular NF-kappaB signaling found in patients with HED-ID.

Amino Acid Sequence↗

PubMed2003

IKK beta plays an essential role in the phosphorylation of RelA/p65 on serine 536 induced by lipopolysaccharide.

Activation of the I kappa B kinase (IKK) complex by LPS induces phosphorylation and degradation of I kappa B alpha, leading to the nuclear translocation of NF-kappa B. Although it is essential for NF-kappa B activation, emerging evidence has indicated that the nuclear translocation of NF-kappa B is not sufficient to activate NF-kappa B-dependent transcription. Here, we reported that LPS induced the phosphorylation of the p65 trans-activation domain on serine 536 in monocytes/macrophages. Using mouse embryonic fibroblasts lacking either IKK alpha or IKK beta, we found that IKK beta played an essential role in LPS-induced p65 phosphorylation on serine 536, while IKK alpha was partially required for the p65 phosphorylation. The LPS-induced p65 phosphorylation on serine 536 was independent of the phosphatidylinositol 3'-kinase/Akt signaling pathway. Furthermore, we found that the phosphorylation on serine 536 increased the p65 transcription activity. In summary, our results demonstrate that IKK beta plays an essential role in the LPS-induced p65 phosphorylation on serine 536, which may represent a mechanism to regulate the NF-kappa B transcription activity by LPS.

Amino Acid Substitution↗

PubMed2003

Telomerase accelerates osteogenesis of bone marrow stromal stem cells by upregulation of CBFA1, osterix, and osteocalcin.

Telomerase activity can prevent telomere shortening and replicative senescence in human somatic cells. We and others have previously demonstrated that forced expression of telomerase in human bone marrow stromal stem cells (BMSSCs) was able to extend their life-span and enhance their bone-forming capability, without inducing malignant transformation. In this study, we determined that telomerase was able to accelerate calcium accumulation of human BMSSCs under osteogenic inductive conditions. Similarly, xenogeneic transplantation of telomerase-expressing BMSSCs (BMSSC-Ts) yielded ectopic bone formation at 2 weeks post-transplantation, 2-4 weeks earlier than typically seen with BMSSCs transfected with empty vector (BMSSC-Cs). Low-density DNA array analysis revealed that telomerase activity increases the expression of G1 regulating genes including cyclin D3, cyclin E1, E2F-4, and DP2, associated with hyperphosphorylation of retinoblastoma (pRb), leading to the extended proliferative capacity of BMSSC-Ts. Importantly, BMSSC-T transplants showed a higher number of human osteogenic cells at 8 weeks post transplantation compared with the BMSSC-C transplants, coupled with a significantly increased osteogenic capacity. One possible mechanism leading to accelerated osteogenesis by BMSSC-Ts may be attributed, at least in part, to the upregulation of the important osteogenic genes such as CBFA1, osterix, and osteocalcin in vitro. Taken together, these findings show that telomerase can accelerate cell cycle progression from G1-to-S phase and enhance osteogenic differentiation of BMSSCs, because of the upregulation of CBFA1, osterix, and osteocalcin.

Alu Elements↗

PubMed2002

Hepatocyte growth factor inhibits anoikis by induction of activator protein 1-dependent cyclooxygenase-2. Implication in head and neck squamous cell carcinoma progression.

Anoikis, also called suspension-induced apoptosis, plays an important role in tumor development, progression, and metastasis. Recently we found that hepatocyte growth factor (HGF) inhibited anoikis of human head and neck squamous cell carcinoma (HNSCC) cells by activating the extracellular signal-regulated kinase (ERK)-signaling pathway. However, the anti-apoptotic effectors that were regulated by the ERK-signaling pathway were unknown. Here we report that HGF-mediated inhibition of anoikis was dependent on activator protein-1 activity through the activation of the ERK-signaling pathway. Using a combination of microarray analysis and Northern blot analysis, we found that an anti-apoptotic gene cyclooxygenase-2 (cox-2) was induced by HGF in an activator protein-1-dependent fashion. Inhibition of Cox-2 activity partially abolished HGF-mediated cell survival, and overexpression of Cox-2 in HNSCC cells provided resistance against anoikis. Moreover, HNSCC cells stably expressing Cox-2 had aggressive tumor growth in a nude mouse model compared with control cells. Taken together, our results demonstrate that Cox-2 plays an important role in HGF-mediated anoikis resistance. HGF may stimulate the progression and growth of HNSCC in vivo by induction of Cox-2.

Animals↗

PubMed2002

c-Myc sensitizes cells to tumor necrosis factor-mediated apoptosis by inhibiting nuclear factor kappa B transactivation.

Nuclear factor kappaB (NF-kappaB) plays a key role in suppression of tumor necrosis factor (TNF)-mediated apoptosis by inducing a variety of anti-apoptotic genes. Expression of c-Myc has been shown to sensitize cells to TNF-mediated apoptosis by inhibiting NF-kappaB activation. However, the precise step in the NF-kappaB signaling pathway and apoptosis modified by c-Myc has not been identified. Using the inducible c-MycER system and c-Myc null fibroblasts, we found that expression of c-Myc inhibited NF-kappaB activation by interfering with RelA/p65 transactivation but not nuclear translocation of NF-kappaB. Activation of c-Myc promoted TNF-induced release of cytochrome c from mitochondria to the cytosol because of the inhibition of NF-kappaB. Furthermore, we found that NF-kappaB-inducible gene A1 was attenuated by expression of c-Myc and that the restoration of A1 expression suppressed c-Myc-induced TNF sensitization. Our results elucidate the molecular mechanisms by which c-Myc increases cell susceptibility to TNF-mediated apoptosis, indicating that c-Myc may exhibit its pro-apoptotic activities by repression of cell survival genes.

Active Transport, Cell Nucleus↗

PubMed2002

Hepatocyte growth factor inhibits anoikis in head and neck squamous cell carcinoma cells by activation of ERK and Akt signaling independent of NFkappa B.

Hepatocyte growth factor (HGF), also known as a scatter factor, regulates a variety of biological activities including cell proliferation, survival, migration, and angiogenesis. Importantly, HGF and its receptor c-Met have been found to be associated with metastasis of human head and neck squamous cell carcinoma (HNSCC). Because anoikis resistance plays an important role in tumor progression and metastasis, here we examined whether HGF suppressed suspension-induced apoptosis (anoikis) in HNSCC cells, and if so, we assessed downstream signaling pathways mediated by HGF. We found that HNSCC cells underwent anoikis upon loss of matrix contact, whereas HGF provided protection against it. HGF-induced anoikis resistance was found to be dependent on both ERK and Akt signaling pathways. The inhibition of either ERK or Akt activation abolished HGF-mediated survival. Furthermore, we found that HGF did not activate NFkappaB transcription in HNSCC cells and that HGF-mediated anoikis resistance was independent of NFkappaB. Taken together, our results suggest that anoikis resistance induced by HGF may also play an important role in the progression and metastasis of HNSCC.

Anoikis↗

PubMed2002

Wnt signaling promotes oncogenic transformation by inhibiting c-Myc-induced apoptosis.

Aberrant activation of the Wnt/beta-catenin signaling pathway is associated with numerous human cancers and often correlates with the overexpression or amplification of the c-myc oncogene. Paradoxical to the cellular transformation potential of c-Myc is its ability to also induce apoptosis. Using an inducible c-MycER expression system, we found that Wnt/beta-catenin signaling suppressed apoptosis by inhibiting c-Myc-induced release of cytochrome c and caspase activation. Both cyclooxygenase 2 and WISP-1 were identified as effectors of the Wnt-mediated antiapoptotic signal. Soft agar assays showed that neither c-Myc nor Wnt-1 alone was sufficient to induce cellular transformation, but that Wnt and c-Myc coordinated in inducing transformation. Furthermore, coexpression of Wnt-1 and c-Myc induced high-frequency and rapid tumor growth in nude mice. Extensive apoptotic bodies were characteristic of c-Myc-induced tumors, but not tumors induced by coactivation of c-Myc and Wnt-1, indicating that the antiapoptotic function of Wnt-1 plays a critical role in the synergetic action between c-Myc and Wnt-1. These results elucidate the molecular mechanisms by which Wnt/beta-catenin inhibits apoptosis and provide new insight into Wnt signaling-mediated oncogenesis.

Animals↗

PubMed2002

Parathyroid hormone and parathyroid hormone-related protein exert both pro- and anti-apoptotic effects in mesenchymal cells.

During bone formation, multipotential mesenchymal cells proliferate and differentiate into osteoblasts, and subsequently many die because of apoptosis. Evidence suggests that the receptor for parathyroid hormone (PTH) and parathyroid hormone-related protein (PTHrP), the PTH-1 receptor (PTH-1R), plays an important role in this process. Multipotential mesenchymal cells (C3H10T1/2) transfected with normal or mutant PTH-1Rs and MC3T3-E1 osteoblastic cells were used to explore the roles of PTH, PTHrP, and the PTH-1R in cell viability relative to osteoblastic differentiation. Overexpression of wild-type PTH-1R increased cell numbers and promoted osteocalcin gene expression versus inactivated mutant receptors. Furthermore, the effects of PTH and PTHrP on apoptosis were dramatically dependent on cell status. In preconfluent C3H10T1/2 and MC3T3-E1 cells, PTH and PTHrP protected against dexamethasone-induced reduction in cell viability, which was dependent on cAMP activation. Conversely, PTH and PTHrP resulted in reduced cell viability in postconfluent cells, which was also dependent on cAMP activation. Further, the proapoptotic-like effects were associated with an inhibition of Akt phosphorylation. These data suggest that parathyroid hormones accelerate turnover of osteoblasts by promoting cell viability early and promoting cell departure from the differentiation program later in their developmental scheme. Both of these actions occur at least in part via the protein kinase A pathway.

3T3 Cells↗

PubMed2002

Bone formation by human postnatal bone marrow stromal stem cells is enhanced by telomerase expression.

Human postnatal bone marrow stromal stem cells (BMSSCs) have a limited life-span and progressively lose their stem cell properties during ex vivo expansion. Here we report that ectopic expression of human telomerase reverse transcriptase (hTERT) in BMSSCs extended their life-span and maintained their osteogenic potential. In xenogenic transplants, hTERT-expressing BMSSCs (BMSSC-Ts) generated more bone tissue, with a mineralized lamellar bone structure and associated marrow, than did control BMSSCs. The enhanced bone-forming ability of BMSSC-Ts was correlated with a higher and sustained expression of the early pre-osteogenic stem cell marker STRO-1, indicating that telomerase expression helped to maintain the osteogenic stem cell pool during ex vivo expansion. These results show that telomerase expression can overcome critical technical barriers to the ex vivo expansion of BMSSCs, and suggest that telomerase therapy may be a useful strategy for bone regeneration and repair.

Adult↗

PubMed2002

The p65/RelA subunit of NF-kappaB suppresses the sustained, antiapoptotic activity of Jun kinase induced by tumor necrosis factor.

Tumor necrosis factor (TNF) signaling through the TNF receptors involves the recruitment of key signaling factors, leading to the activation of both the transcription factor NF-kappaB and the stress-activated Jun kinase (JNK). In most cells, TNF signaling leads to a rapid and transient increase in JNK activity. However, we show that TNF treatment leads to the sustained activation of JNK in cells that are null for the p65/RelA subunit of NF-kappaB as well as in cells expressing the super-repressor form of IkappaB. In addition, the data indicate that the ability of p65/RelA to regulate gene expression is required to suppress the persistent activation of JNK. Interestingly, this suppression occurs upstream of JNK, within the signal transduction cascade leading to JNK activation, without affecting the stress-activated kinase p38. Since NF-kappaB has previously been shown to be involved in the suppression of TNF-induced apoptosis, we were interested in determining the role of deregulated JNK activity, induced by the loss of NF-kappaB, in controlling the cell death response. Through the use of different approaches for inhibition of JNK, we show that the suppression of JNK activity in cells that lack active NF-kappaB enhances the apoptotic response to TNF. These data suggest that the activity of JNK in cells blocked for NF-kappaB function provides an antiapoptotic signal and explains, at least partly, why a significant number of NF-kappaB null cells remain viable following TNF treatment.

Animals↗

PubMedDate not supplied

Investigation of multipotent postnatal stem cells from human periodontal ligament.

BACKGROUND: Periodontal diseases that lead to the destruction of periodontal tissues--including periodontal ligament (PDL), cementum, and bone--are a major cause of tooth loss in adults and are a substantial public-health burden worldwide. PDL is a specialised connective tissue that connects cementum and alveolar bone to maintain and support teeth in situ and preserve tissue homoeostasis. We investigated the notion that human PDL contains stem cells that could be used to regenerate periodontal tissue. METHODS: PDL tissue was obtained from 25 surgically extracted human third molars and used to isolate PDL stem cells (PDLSCs) by single-colony selection and magnetic activated cell sorting. Immunohistochemical staining, RT-PCR, and northern and western blot analyses were used to identify putative stem-cell markers. Human PDLSCs were transplanted into immunocompromised mice (n=12) and rats (n=6) to assess capacity for tissue regeneration and periodontal repair. Findings PDLSCs expressed the mesenchymal stem-cell markers STRO-1 and CD146/MUC18. Under defined culture conditions, PDLSCs differentiated into cementoblast-like cells, adipocytes, and collagen-forming cells. When transplanted into immunocompromised rodents, PDLSCs showed the capacity to generate a cementum/PDL-like structure and contribute to periodontal tissue repair. INTERPRETATION: Our findings suggest that PDL contains stem cells that have the potential to generate cementum/PDL-like tissue in vivo. Transplantation of these cells, which can be obtained from an easily accessible tissue resource and expanded ex vivo, might hold promise as a therapeutic approach for reconstruction of tissues destroyed by periodontal diseases.

Adult↗