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Suwon Kim - One of the best experts on this subject based on the ideXlab platform.

  • Abstract 3453: CXCL10 contributes to aggressive disease progression in ING4-deficient breast cancer
    Molecular and Cellular Biology Genetics, 2019
    Co-Authors: Emily Szeto, Suwon Kim
    Abstract:

    CXCL10 is a chemoattractant secreted by various cell types during inflammation for immune cells expressing its cognate receptor CXCR3. In breast cancer, CXCL10 has been associated with increased tumor lymphocytic infiltrates and poor patient prognosis. In addition, pharmacological inhibition of CXCR3 resulted in reduced lung metastases of mammary tumors in mice, suggesting CXCL10/CXCR3 signaling contributes to aggressive breast cancer. We previously identified CXCL10 as an NF-κB target gene repressed by Inhibitor of Growth 4 (ING4) in breast cancer cells. As ING4 deficiencies reported in 34% of breast tumors have been correlated with faster disease recurrence in patients, we investigated a functional interplay between CXCL10 and ING4. First, we assessed the clinical relevance of CXCL10, CXCR3, and/or ING4 expression levels using a public gene expression data set. Second, we genetically engineered T47D breast cancer cells to overexpress or delete the ING4 gene and determined cell phenotypes in the presence or absence of CXCL10 by utilizing cell migration assays and Western blot. Analysis of the GDS806 gene data set showed that patients with tumors expressing high levels of CXCL10 experienced significantly increased rates of disease recurrence while CXCR3 expression did not have a significant effect. The analysis also showed tumors expressing low levels of ING4 expressed higher levels of CXCL10, indicating an inverse expression pattern between the two genes. Moreover, patients with ING4-low/CXCL10-high tumors had a 4-fold faster recurrence rate compared to ING4-high/CXCL10-low tumor patients. These results indicated that high CXCL10 expression contributed to significantly accelerated disease recurrence compounded by low ING4 expression, suggesting a functional relationship between the two genes which may exacerbate tumor phenotypes. While CXCL10 did not affect growth rates of T47D cells with or without ING4, CXCL10 induced migration only in ING4-deleted cells. These results suggested that ING4 inhibited CXCL10 signaling. The western blot analyses showed no change in CXCR3 expression, but elevated phosphorylation of insulin-like growth factor 1 receptor (IGF1R) after CXCL10 treatment, suggesting CXCL10 may signal by activating IGF1R. Consistent with this idea, when cells were treated with an IGF1R inhibitor, Linsitinib (OSI-906), CXCL10-induced migration was attenuated in ING4-deleted cells. In conclusion, these results suggest that in order to promote an aggressive tumor phenotype, CXCL10 induces migration of ING4-deficient tumor cells in part by activating IGF1R. This study reports the first demonstration of the chemokine CXCL10 exerting a direct effect on breast cancer cells and puts forth the CXCL10/IGF1R signaling pathway as a potential therapeutic target for ING4-deficient aggressive breast cancer. Citation Format: Emily Szeto, Suwon Kim. CXCL10 contributes to aggressive disease progression in ING4-deficient breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3453.

  • Tumor suppressor ING4 inhibits estrogen receptor activity in breast cancer cells
    Breast cancer (Dove Medical Press), 2016
    Co-Authors: Madeline M. Keenen, Suwon Kim
    Abstract:

    Abstract Resistance to antiestrogen therapy remains a significant problem in breast cancer. Low expression of inhibitor of growth 4 (ING4) in primary tumors has been correlated with increased rates of recurrence in estrogen receptor-positive (ER+) breast cancer patients, suggesting a role for ING4 in ER signaling. This study provides evidence that ING4 inhibits ER activity. ING4 overexpression increased the sensitivity of T47D and MCF7 ER+ breast cancer cells to hormone deprivation. ING4 attenuated maximal estrogen-dependent cell growth without affecting the dose-response of estrogen. These results indicated that ING4 functions as a noncompetitive inhibitor of estrogen signaling and may inhibit estrogen-independent ER activity. Supportive of this, treatment with fulvestrant but not tamoxifen rendered T47D cells sensitive to hormone deprivation as did ING4 overexpression. ING4 did not affect nuclear ERα protein expression, but repressed selective ER-target gene transcription. Taken together, these results demonstrated that ING4 inhibited estrogen-independent ER activity, suggesting that ING4-low breast tumors recur faster due to estrogen-independent ER activity that renders tamoxifen less effective. This study puts forth fulvestrant as a proposed therapy choice for patients with ING4-low ER+ breast tumors.

  • Transient Induction of ING4 by Myc Drives Prostate Epithelial Cell Differentiation and Its Disruption Drives Prostate Tumorigenesis
    Cancer research, 2014
    Co-Authors: Penny L. Berger, Suwon Kim, Sander B. Frank, Veronique V. Schulz, Eric A. Nollet, Mathew J. Edick, Brittany Holly, Ting Tung A. Chang, Galen Hostetter, Cindy K. Miranti
    Abstract:

    The mechanisms by which Myc overexpression or Pten loss promotes prostate cancer development are poorly understood. We identified the chromatin remodeling protein, ING4, as a crucial switch downstream of Myc and Pten that is required for human prostate epithelial differentiation. Myc-induced transient expression of ING4 is required for the differentiation of basal epithelial cells into luminal cells, while sustained ING4 expression induces apoptosis. ING4 expression is lost in >60% of human primary prostate tumors. ING4 or Pten loss prevents epithelial cell differentiation, which was necessary for tumorigenesis. Pten loss prevents differentiation by blocking ING4 expression, which is rescued by ING4 re-expression. Pten or ING4 loss generates tumor cells that co-express basal and luminal markers, indicating prostate oncogenesis occurs through disruption of an intermediate step in the prostate epithelial differentiation program. Thus, we identified a new epithelial cell differentiation switch involving Myc, Pten, and ING4, which when disrupted leads to prostate tumorigenesis. Myc overexpression and Pten loss are common genetic abnormalities in prostate cancer, whereas loss of the tumor suppressor ING4 has not been reported. This is the first demonstration that transient ING4 expression is absolutely required for epithelial differentiation, its expression is dependent on Myc and Pten, and it is lost in the majority of human prostate cancers. This is the first demonstration that loss of ING4, either directly or indirectly through loss of Pten, promotes Myc-driven oncogenesis by deregulating differentiation. The clinical implication is that Pten/ING4 negative and ING4-only negative tumors may reflect two distinct subtypes of prostate cancer.

  • Negative regulation of NF-κB by the ING4 tumor suppressor in breast cancer.
    PloS one, 2012
    Co-Authors: Sara A. Byron, Coya Tapia, Galen Hostetter, Elizabeth Min, Tanya S. Thal, Aprill Watanabe, David O. Azorsa, Tanya H. Little, Suwon Kim
    Abstract:

    Nuclear Factor kappa B (NF-κB) is a key mediator of normal immune response but contributes to aggressive cancer cell phenotypes when aberrantly activated. Here we present evidence that the Inhibitor of Growth 4 (ING4) tumor suppressor negatively regulates NF-κB in breast cancer. We surveyed primary breast tumor samples for ING4 protein expression using tissue microarrays and a newly generated antibody. We found that 34% of tumors expressed undetectable to low levels of the ING4 protein (n = 227). Tumors with low ING4 expression were frequently large in size, high grade, and lymph node positive, suggesting that down-regulation of ING4 may contribute to breast cancer progression. In the same tumor set, we found that low ING4 expression correlated with high levels of nuclear phosphorylated p65/RelA (p-p65), an activated form of NF-κB (p = 0.018). Fifty seven percent of ING4-low/p-p65-high tumors were lymph node-positive, indicating a high metastatic tendency of these tumors. Conversely, ectopic expression of ING4 inhibited p65/RelA phosphorylation in T47D and MCF7 breast cancer cells. In addition, ING4 suppressed PMA-induced cell invasion and NF-κB-target gene expression in T47D cells, indicating that ING4 inhibited NF-κB activity in breast cancer cells. Supportive of the ING4 function in the regulation of NF-κB-target gene expression, we found that ING4 expression levels inversely correlated with the expression of NF-κB-target genes in primary breast tumors by analyzing public gene expression datasets. Moreover, low ING4 expression or high expression of the gene signature composed of a subset of ING4-repressed NF-κB-target genes was associated with reduced disease-free survival in breast cancer patients. Taken together, we conclude that ING4 negatively regulates NF-κB in breast cancer. Consequently, down-regulation of ING4 leads to activation of NF-κB, contributing to tumor progression and reduced disease-free patient survival in breast cancer.

  • deletion of the inhibitor of growth 4 ING4 tumor suppressor gene is prevalent in human epidermal growth factor 2 her2 positive breast cancer
    Human Pathology, 2011
    Co-Authors: Suwon Kim, Coya Tapia, Inti Zlobec, Sandra Schneider, Ergin Kilic, Uwe Guth, Lukas Bubendorf
    Abstract:

    Inhibitor of growth 4 (ING4) is a candidate tumor suppressor gene that was shown to be deleted in 10% to 20% of breast cancers by array comparative genome hybridization analysis. We developed fluorescent in situ hybridization to detect the ING4 gene directly in the tissue samples on tumor tissue microarrays. We evaluated the ING4 gene status in 1033 breast cancer tissue samples and observed that ING4 was deleted in 16.5% (170/1033) of all breast cancers. ING4 deletion was significantly associated with Her2 overexpression: of the tumors with ING4 deletion, 23.8% (39/164) were human epidermal growth factor 2 (HER2) positive, as compared with 14.1% (115/814) of the tumors without ING4 deletion (P = .002). In addition, the tumors with ING4 deletion were more likely to belong to the HER2 molecular subtype (estrogen receptor negative/progesterone receptor negative/human epidermal growth factor positive) of breast cancer, compared with the other subtypes (28.4% HER2 versus 15.7% all, P = .002). ING4 deletion did not affect survival outcome of all patients with breast cancer (P = .797) or of the patients with HER2-positive tumors (P = .792). We conclude that ING4 deletion in breast cancer is relatively common, as 1 in 6 breast cancer harbors ING4 deletion. Furthermore, ING4 deletion is more prevalent in HER2-positive tumors, suggesting a functional antagonistic relationship between the ING4 tumor suppressor and the HER2 oncogene. These results sustain the view that ING4 is a tumor suppressor in breast cancer and suggest that ING4 deletion may contribute to the pathogenesis of HER2-positive breast cancer.

Rémy Pedeux - One of the best experts on this subject based on the ideXlab platform.

  • Human ex vivo prostate tissue model system identifies ING3 as an oncoprotein
    British Journal of Cancer, 2018
    Co-Authors: Urszula L Mcclurg, Arash Nabbi, Charles Ricordel, Svitlana Korolchuk, Stuart Mccracken, Rakesh Heer, Laura Wilson, Lisa M Butler, Bronwyn Kate Irving-hooper, Rémy Pedeux
    Abstract:

    Background: Although the founding members of the INhibitor of Growth (ING) family of histone mark readers, ING1 and ING2, were defined as tumour suppressors in animal models, the role of other ING proteins in cellular proliferation and cancer progression is unclear. Methods: We transduced ex vivo benign prostate hyperplasia tissues with inducible lentiviral particles to express ING proteins. Proliferation was assessed by H3S10^phos immunohistochemistry (IHC). The expression of ING3 was assessed by IHC on a human prostate cancer tissue microarray (TMA). Gene expression was measured by DNA microarray and validated by real-time qPCR. Results: We found that ING3 stimulates cellular proliferation in ex vivo tissues, suggesting that ING3 could be oncogenic. Indeed, ING3 overexpression transformed normal human dermal fibroblasts. We observed elevated levels of ING3 in prostate cancer samples, which correlated with poorer patient survival. Consistent with an oncogenic role, gene-silencing experiments revealed that ING3 is required for the proliferation of breast, ovarian, and prostate cancer cells. Finally, ING3 controls the expression of an intricate network of cell cycle genes by associating with chromatin modifiers and the H3K4^me3 mark at transcriptional start sites. Conclusions: Our investigations create a shift in the prevailing view that ING proteins are tumour suppressors and redefine ING3 as an oncoprotein.

  • ING1 and ING2: multifaceted tumor suppressor genes
    Cellular and Molecular Life Sciences, 2013
    Co-Authors: Claire Guérillon, Delphine Larrieu, Rémy Pedeux
    Abstract:

    In hibitor of G rowth 1 ( ING1 ) was identified and characterized as a “candidate” tumor suppressor gene in 1996. Subsequently, four more genes, also characterized as “candidate” tumor suppressor genes, were identified by homology search: ING2, ING3, ING4 , and ING5. The ING proteins are characterized by a high homology in their C-terminal domain, which contains a Nuclear Localization Sequence and a Plant HomeoDomain (PHD), which has a high affinity to Histone 3 tri-methylated on lysine 4 (H3K4Me3). The ING proteins have been involved in the control of cell growth, senescence, apoptosis, chromatin remodeling, and DNA repair. Within the ING family, ING1 and ING2 form a subgroup since they are evolutionarily and functionally close. In yeast, only one gene, Pho23, is related to ING1 and ING2 and possesses also a PHD. Recently, the ING1 and ING2 tumor suppressor status has been fully established since several studies have described the loss of ING1 and ING2 protein expression in human tumors and both ING1 and ING2 knockout mice were reported to have spontaneously developed tumors, B cell lymphomas, and soft tissue sarcomas, respectively. In this review, we will describe for the first time what is known about the ING1 and ING2 genes, proteins, their regulations in both human and mice, and their status in human tumors. Furthermore, we explore the current knowledge about identified functions involving ING1 and ING2 in tumor suppression pathways especially in the control of cell cycle and in genome stability.

  • ING1 and ING2: Multi-Faceted Tumor Suppressor Genes
    Cellular and Molecular Life Sciences, 2013
    Co-Authors: Claire Guérillon, Delphine Larrieu, Rémy Pedeux
    Abstract:

    ING1 (Inhibitor of Growth 1) was identified and characterized as a "candidate" tumor suppressor gene in 1996. Subsequently four more genes, also characterized as "candidate" tumor suppressor genes, were identified by homology search: ING2, ING3, ING4 and ING5. The ING proteins are characterized by a high homology in their C-terminal domain which contains a Nuclear Localization Sequence (NLS) and a Plant HomeoDomain (PHD) which has a high affinity to Histone 3 tri-methylated on lysine 4 (H3K4Me3). The ING proteins have been involved in the control of cell growth, senescence, apoptosis, chromatin remodelling and DNA repair. Within the ING family, ING1 and ING2 form a subgroup since they are evolutionarily and functionally close. In Yeast, only one gene, Pho23, is related to ING1 and ING2 and possesses also a PHD. Recently, the ING1 and ING2 tumor suppressor status has been fully established since several studies have described the loss of ING1 and ING2 protein expression in human tumors and both ING1 and ING2 knockout mice were reported to have spontaneously developed tumors, B-cell lymphomas and soft tissue sarcomas respectively. In this review we will describe for the first time what is known about ING1 and ING2 genes, proteins, their regulations in both human and mice, and their status in human tumors. Furthermore, we explore the current knowledge about identified functions involving ING1 and ING2 in tumor suppression pathways especially in the control of cell cycle and in genome stability.

  • Ing2 sumoylation regulates cell proliferation
    Cancer Research, 2008
    Co-Authors: Damien Ythier, Delphine Larrieu, Romuald Binet, Christian Brambilla, Damien Nissou, Elisabeth Brambilla, Rémy Pedeux
    Abstract:

    910 ING genes (Inhibitor of Growth, ING1-5) are candidate tumor suppressor genes which have been recently identified and characterized. ING proteins contain a very highly conserved C-terminus region in which a Nuclear Localization Sequence (NLS) and a Plant HomeoDomain (PHD) are present. PHD fingers are commonly found in chromatin-regulatory proteins. All ING proteins are components of Histone AcetylTransferase (HAT) or Histone DeACetylase (HDAC) complexes. Regarding ING2, it is a stable component of the Sin3A/HDAC1 complex. The PHD of ING2 has a high affinity for the tail of histone 3 tri-methylated on the lysine 4. This interaction allows the recruitment of the Sin3A-HDAC chromatin remodeling complex to the promoter of proliferative genes such as cyclin D1 and c-myc.
 Immunoprecipitation of Sin3A resulted in the detection of ING2 but in addition a second band about 10 kDa higher was detected. It suggested that ING2 may be post-translationally modified. In-vitro SUMOylation experiments performed on GST-ING2 showed that ING2 can be SUMOylated on at least two residues. We confirmed by co-transfection experiments of ING2 and SUMO in U2OS cells that ING2 can also be SUMOylated in-vivo . Using truncated constructs of ING2 we have zeroed-in the region of ING2 that is SUMOylated. In parallel, a bioinformatics analysis of the amino acid sequence of ING2 revealed the presence of several known consensus sites for SUMOylation (ΨKXE). These experiments allowed us to identify two potential sites for SUMOylation. Directed mutagenesis on these two candidate SUMOylated residues were performed, allowing the identification of one lysine residue strongly SUMOylated in-vivo . Subsequently, we showed that ING2 SUMOylation did not affect its nuclear localization but was important for ING2 binding to the Sin3A complex. Taking into account the recently published data involving ING2 in the recruitment of the Sin3A-HDAC complex to the promoter of proliferative genes, we have tested the involvement of ING2 SUMOylation on the repression of proliferative genes. Cell growth experiments (growth curve and colony formation assays) revealed that ING2 SUMOylated mutant was not able to repress cell proliferation as wild-type ING2 did.
 Taken together our results show that the candidate tumor suppressor protein ING2 is SUMOylated and that this SUMOylation is necessary for ING2 binding with Sin3A complexes. Moreover, SUMOylation of ING2 appears to be essential for repressing cell proliferation. Thus, our results suggest that SUMOylation could regulate ING2 ability to recruit Sin3A-HDAC complex to the chromatin. Therefore, SUMOylation of ING2 would be critical to regulate the expression of genes involved in cell cycle control.

  • Inhibitor of growth 4 suppresses cell spreading and cell migration by interacting with a novel binding partner, liprin alpha1.
    Cancer Research, 2007
    Co-Authors: Jiang Cheng Shen, Damien Ythier, Rémy Pedeux, Kensuke Kumamoto, Motoko Unoki, Lyuba Varticovski, Alain Duperray, Curtis Harris
    Abstract:

    Inhibitor of growth 4 (ING4) is a candidate tumor suppressor that plays a major role in gene regulation, cell cycle control, apoptosis, and angiogenesis. ING4 expression is down-regulated in glioblastoma cells and head and neck squamous cell carcinoma. Here, we identified liprin alpha1/PPFIA1, a cytoplasmic protein necessary for focal adhesion formation and axon guidance, as a novel interacting protein with ING4. ING4 and liprin alpha1 colocalized at lamellipodia in the vicinity of vinculin. Overexpressed ING4 suppressed cell spreading and cell migration. In contrast, overexpressed liprin alpha1 enhanced cell spreading and cell migration. Knockdown of endogenous ING4 with RNA interference induced cell motility, whereas knockdown of endogenous liprin alpha1 suppressed cell motility. ING4 also suppressed cell motility that was enhanced by liprin alpha1. However, ING4 did not further suppress cell motility when liprin alpha1 was suppressed with RNA interference, suggesting a functional and mechanistic interdependence between these proteins. In addition to its nuclear functions, cytoplasmic ING4 interacts with liprin alpha1 to regulate cell migration and, with its known antiangiogenic function, may prevent invasion and metastasis.

Ji-cheng Yang - One of the best experts on this subject based on the ideXlab platform.

  • adenovirus mediated ING4 pten double tumor suppressor gene co transfer modified by rgd enhances antitumor activity in human nasopharyngeal carcinoma cells
    International Journal of Oncology, 2015
    Co-Authors: Yihong Wang, Yu-feng Xie, Ji-cheng Yang, Weihua Sheng, Jisheng Liu
    Abstract:

    Inhibitor of growth-4 (ING4) is a member of the inhibitor of growth (ING) family and acts as a tumor suppressor protein. PTEN is a phosphatase and shows potent and extensive antitumor activity. In this study, we constructed an RGD-modified bicistronic ING4/PTEN adenovirus (Ad.RGD-ING4-PTEN) and comprehensively investigated its effects following modification of the CNE human nasopharyngeal carcinoma cell line both in vitro and in vivo. We demonstrated that Ad.RGD-ING4-PTEN enhanced growth inhibition and apoptosis. Furthermore, expression of P21, Bax and cleaved caspase-3 was upregulated, while that of Bcl-2 and survivin was downregulated in CNE cells and CNE xenografted tumors. Moreover, Ad.RGD-ING4-PTEN treatment additively downregulated CD34, VEGF and microvessel density in subcutaneously (s.c.) xenografted CNE cell tumors. The enhanced antitumor activity generated by Ad.RGD-ING4-PTEN was closely associated with activation of the intrinsic and extrinsic apoptotic pathways and additive inhibition of tumor angiogenesis both in vitro and in vivo. On the basis of this evidence, it is believed that cancer gene therapy combining two tumor suppressors such as ING4 and PTEN can be used to establish an effective and novel therapeutic strategy for nasopharyngeal carcinoma and other cancers.

  • Adenovirus-mediated ING4 Gene Transfer in Osteosarcoma Suppresses Tumor Growth via Induction of Apoptosis and Inhibition of Tumor Angiogenesis.
    Technology in cancer research & treatment, 2014
    Co-Authors: Yu-feng Xie, Jing-cheng Miao, Weihua Sheng, Ji-cheng Yang
    Abstract:

    The inhibitor of growth (ING) family proteins have been defined as candidate tumor suppressors. ING4 as a novel member of ING family has potential tumor-suppressive effects via multiple pathways. However, the therapeutic effect of adenovirus-mediated ING4 (Ad-ING4) gene transfer in human osteosarcoma is still unknown. In this study, we explored the in vitro and in vivo antitumor activity of Ad-ING4 in human osteosarcoma and its potential mechanism using a MG-63 human osteosarcoma cell line. We demonstrated that Ad-ING4 induced significant growth inhibition and apoptosis, upregulated the expression of P21, P27 and Bax, downregulated the Bcl-2 expression and activated Caspase-3 in MG-63 human osteosarcoma cells. Moreover, intratumoral injections of Ad-ING4 in athymic nude mice bearing MG-63 human osteosarcoma tumors significantly suppressed osteosarcoma xenografted tumor growth, increased the expression of P21, P27 and Bax, reduced the Bcl-2 and CD34 expression and microvessel density (MVD) in tumors. This ...

  • Synergistic tumor suppression by Ad. RGD-ING4 in human nasopharyngeal carcinoma cell CNE and its mechanism
    Lin chuang er bi yan hou tou jing wai ke za zhi = Journal of clinical otorhinolaryngology head and neck surgery, 2014
    Co-Authors: Yihong Wang, Ji-cheng Yang, Weihua Sheng, Jisheng Liu
    Abstract:

    OBJECTIVE To investigate the effect of adenovirus-mediated ING4 with RGD on proliferation, apoptosis and cell cycle of human nasopharyngeal carcinoma cell CNE and explore its probable mechanism. METHOD CNE cells were infected with Ad. RGD-ING4 and adenovirus vector, ING4 gene expression level was detected by RT-PCR and the target protein expression was tested by Western blot. MTT assay was adopted to evaluate the efect of ING4 on cell growth of CNE, Annexin -V-PE/7-AAD Double staining was used to measure the efect of ING4 on apoptosis, and PI staining was used to measure the efect of ING4 on the cell cycle. Differential expression of P21, Bcl-2 and Bax gene was detected by RT-PCR,and Differential expression of Survivin and Caspase 3 protein was detected by Western blot. RESULT CNE cells were cultured with Ad. RGD-ING4 for 72 h ,the results showed that ING4 was overexpressed in CNE cells ,the growth of CNE cells was obviously inhibited , apoptosis rate was significantly increased and G2/M phase was arrested apparently. The results of RT-PCR showed that Ad. RGD-ING4 significantly down-regulated the Bcl-2 and up-regulates the Bax and P21 expression in CNE cells, and the difference was statistically significant(P < 0.01). Western blot showed that the expression of Survivin was decreased and Cleaved-Caspase 3 was increased. CONCLUSION Ad. RGD-ING4 can play the role of tumor suppressor synergies on nasopharyngeal carcinoma cell CNE by down-regulating Bcl-2, Survivin expression and up-regulating P21, Bax and Cleaved-Caspase 3 expression.

  • Autophagy contributes to ING4-induced glioma cell death.
    Experimental cell research, 2013
    Co-Authors: Aihua Gong, Yan Zhang, Er-meng Xiong, Wenjie Guo, Wanxin Peng, Genbao Shao, Jie Jin, Zhijian Zhang, Ji-cheng Yang
    Abstract:

    Previous studies suggest that ING4, a novel member of ING (inhibitor of growth) family, can inhibit brain tumor growth. However, whether autophagy is involved in ING4-induced cell death still remains unknown. In this study, we found that in addition to apoptosis, autophagy also contributed to cell death induced by ING4. Autophagy levels were elevated following the exposure to Ad-ING4, including enhanced fluorescence intensity of monodansylcadervarine (MDC), a specific in vivo marker for autophagic vacuoles, and increased expression levels of the LC3-II and Beclin-1, wheras the autophagic levels were attenuated following the pretreatment of 3-MA, the inhibitor of autophagy, which significantly decreased the Ad-ING4-induced cell death compared with caspase inhibitor zVAD. Furthermore, ING4 also induced mitochondrial dysfunction, such as mitophagy, collapse of mitochondrial membrane potential and the intracellular ROS, which indicated that mitochondria might be associated with the process of autophagic cell death of glioma cells. Finally, the relationship among Bax, Bcl-2, Beclin-1 and caspase family proteins levels were analyzed in glioma cells U251MG and LN229 infected with Ad-ING4 or Ad-lacZ. It is suggested that both autophagy and apoptosis could contribute to ING4-induced glioma cell death, and mitochondria might play an important role in this process. Our findings reveal novel aspects of the autophagy in glioma cells that underlie the cytotoxic action of ING4, possibly providing new insights in the development of combinatorial therapies for gliomas.

  • enhanced tumor suppression by an ING4 il 24 bicistronic adenovirus mediated gene cotransfer in human non small cell lung cancer cells
    Cancer Gene Therapy, 2011
    Co-Authors: Y Zhu, Yu-feng Xie, Jim Xiang, W Sheng, Ji-cheng Yang
    Abstract:

    ING4 as a member of inhibitor of growth (ING) tumor suppressor family has potent inhibitory effects on a variety of tumors. Interleukin-24 (IL-24), a cytokine-tumor suppressor, also shows broad-spectrum and tumor-specific antitumor activities. In this report, we constructed an ING4/IL-24 bicistronic adenovirus (Ad-ING4-IL-24) and assessed its combined effect on in vitro and in vivo A549 human non-small cell lung cancer cells. We demonstrated that ING4 and IL-24 combination treatment by adenovirus-mediated ING4 and IL-24 coexpression induced additive growth suppression and apoptosis as well as an overlapping effect on upregulation of P21, P27, Fas, Bax and cleaved Caspases-8, 9, 3 and downregulation of Bcl-2 in in vitro A549 lung carcinoma cells. Moreover, Ad-ING4-IL-24 treatment additively inhibited in vivo A549 lung carcinoma subcutaneous (s.c.) xenografted tumor growth and reduced CD34 and microvessel density in A549 xenografted tumors in athymic nude mice. The enhanced antitumor activity elicited by Ad-ING4-IL-24 was closely associated with the coordinate activation of extrinsic and intrinsic apoptotic pathways and additive inhibition of tumor angiogenesis. Thus, our results indicate that cancer gene therapy combining two or more tumor suppressors such as ING4 and IL-24 may constitute a novel and effective therapeutic strategy for lung carcinoma and other cancers.

Curtis C. Harris - One of the best experts on this subject based on the ideXlab platform.

  • Regulation of cell spreading and migration by candidate tumor suppressor ING4
    Cancer Research, 2006
    Co-Authors: Jiang C. Shen, Rémy Pedeux, Kensuke Kumamoto, Motoko Unoki, Lyuba Varticovski, Curtis C. Harris
    Abstract:

    2484 Jiang Cheng Shen 1 , Kensuke Kumamoto 1 , Motoko Unoki 1 , Lyuba Varticovski 1 , Remy Pedeux 2 , and Curtis C. Harris 1 1 Laboratory of Human Carcinogenesis, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA, and 2 INSERM U578, Institut Albert Bonniot, France The candidate tumor suppressor ING4 belongs to a gene family initially manifested in inhibition of cell growth involving in p53-dependent gene regulation pathways. Recent studies further suggested the ING4 function in regulation of angiogenesis induced by human glioblastoma cells through the NF-κB pathway; and in suppression of lost of contact inhibition by ectopic expression of human ING4 in myc -elicited rat cells. Thus ING4 may play a role in modulation of cell morphology and/or cell motility. Here we report that ING4 regulates cell spreading and migration. We first demonstrated that expression of ING4 in RKO cells suppresses cell migration in modified Boyden chamber and siRNA knock-down of ING4 enhances the migration. Observation of single cell spreading by phalloidin staining on F-actin also indicated that ING4 modulates actin filament organization and thus slows down cell spreading, which is consistent with results from monolayer wound-healing assay that ING4 expression in RKO cells retards the gap-filling capability after inflicting a scratch wound on the cell monolayer. Cytoplasmic localization of ING4 can be visualized by immunofluorescence microscopy and ING4 proteins are accumulating in the protruding lamellipodium, suggesting a role for ING4 in cytoskeleton organization during cell spreading. To understand the network of ING4-interacting proteins, we carried out protein affinity pull-down and by using mass spectrometry we identified a target protein, liprin α1, which plays an important role in regulation of focal adhesion. Subsequent co-immunoprecipitation of ING4 and liprin α1 further confirmed this physical interaction, by which we suggest a possible role of ING4 in focal adhesion regulation. Further analyses by immunofluorescence showed that ING4 either co-localizes with, or proximately localizes to, the components of focal adhesion complex during cell spreading. We thus propose that ING4 involves in regulation of focal adhesion during cell spreading and migration. In summary, we have demonstrated that ING4 regulates cell spreading and migration. The cytoplasmic functions of ING4, including accumulation in lamellipodium, co-localization with focal adhesion complex, and interaction with liprin α1, have provided molecular evidence to substantiate this observation. Our results therefore shed a new light on studies for ING4 function in signal transduction and tumor suppression.

  • Identification of three novel splice-variants of a tumor suppressor candidate, ING4
    Cancer Research, 2006
    Co-Authors: Motoko Unoki, Jiang Cheng Shen, Zheng Zhi-ming, Curtis C. Harris
    Abstract:

    Proc Amer Assoc Cancer Res, Volume 47, 2006 2485 The inhibitor of growth family member 4 (ING4) was identified in our laboratory as a candidate tumor suppressor gene that has growth suppressive effect. It has been also reported from other laboratories that the gene is down regulated in glioblastoma cells and head and neck squamous cell carcinoma. Here we identified three novel splice variants of ING4, ING4\_v2 (3 bp skip type), ING4\_v3 (9 bp skip type), and ING4\_v4 (12 bp skip type), that are produced from a combination of two splice donor sites at the end of exon 4 and two splice acceptor sites at the start of exon 5 of the ING4 gene. The longest splice variant (ING4\_v1) that was originally enrolled as ING4 encodes an intact NLS that is important for p53 binding besides nuclear localization, while the other three splice variants partially lack NLS, resulting in an increase of cytoplasmic localization. Recently, although it is known that ING4 has a variety of functions in cells, the entire role of ING4 has not been elucidated. We found one of the three NLS lacking ING4 variants, ING4\_v2, was expressed in cells at almost the same level of the original longest type, suggesting ING4 also has an important role in cytoplasm. Growth suppressive effect of the variants that have the partial NLS (ING4\_v2 and ING4\_v4) was attenuated by weaker effect of the variants on p21 promoter activation. We also found that the ING4\_v4 acts as a dominant negative on induction of p21 by ING4_v1. The functional roles of the proteins generated by alternate spliced forms of ING4 may provide new insight into human carcinogenesis.

Francisco J Blanco - One of the best experts on this subject based on the ideXlab platform.

  • crystal structure of inhibitor of growth 4 ING4 dimerization domain reveals functional organization of ing family of chromatin binding proteins
    Journal of Biological Chemistry, 2012
    Co-Authors: Simone Culurgioni, Alicia Palacios, Francisco J Blanco, Ines G Munoz, Guillermo Montoya, Alberto Moreno, Ignacio Palmero, Maider Villate
    Abstract:

    The protein ING4 binds to histone H3 trimethylated at Lys-4 (H3K4me3) through its C-terminal plant homeodomain, thus recruiting the HBO1 histone acetyltransferase complex to target promoters. The structure of the plant homeodomain finger bound to an H3K4me3 peptide has been described, as well as the disorder and flexibility in the ING4 central region. We report the crystal structure of the ING4 N-terminal domain, which shows an antiparallel coiled-coil homodimer with each protomer folded into a helix-loop-helix structure. This arrangement suggests that ING4 can bind simultaneously two histone tails on the same or different nucleosomes. Dimerization has a direct impact on ING4 tumor suppressor activity because monomeric mutants lose the ability to induce apoptosis after genotoxic stress. Homology modeling based on the ING4 structure suggests that other ING dimers may also exist.

  • Functional impact of cancer-associated mutations in the tumor suppressor protein ING4.
    Carcinogenesis, 2010
    Co-Authors: Alberto Moreno, Alicia Palacios, Francisco J Blanco, Jose L. Orgaz, Benilde Jiménez, Ignacio Palmero
    Abstract:

    Inhibitor of growth 4 (ING4) is a member of the ING family of tumor suppressor proteins. In this study, we have analyzed the impact of two mutations in ING4 associated with human tumors (Y121N and N214D), testing their behavior in a series of functional, biochemical and structural analyses. We report that the N214D mutation dramatically dampened the ability of ING4 to inhibit proliferation, anchorage-independent growth or cell migration or to sensitize to cell death. In turn, the Y121N mutant did not differ significantly from wild-type ING4 in our assays. Neither of the mutations altered the normal subcellular localization of ING4, showing predominantly nuclear accumulation. We investigated the molecular basis of the defect in the activity of the N214D mutant. The folding and ability to bind histone marks of ING4 was not significantly altered by this mutation. Instead, we found that the functional impairment of the N214D mutant correlates with reduced protein stability due to increased proteasome-mediated degradation. In summary, our data demonstrates that a point mutation of ING4 associated to human tumors leads to the loss of several essential functions of ING4 pertinent to tumor protection and highlight the importance of ING4 function to prevent tumorigenesis.

  • crystallization and preliminary x ray diffraction analysis of the dimerization domain of the tumour suppressor ING4
    Acta Crystallographica Section F-structural Biology and Crystallization Communications, 2010
    Co-Authors: Simone Culurgioni, Alicia Palacios, Francisco J Blanco, Ines G Munoz, Pilar Redondo, Guillermo Montoya
    Abstract:

    Inhibitor of growth protein 4 (ING4) belongs to the ING family of tumour suppressors and is involved in chromatin remodelling, in growth arrest and, in cooperation with p53, in senescence and apoptosis. Whereas the structure and histone H3-binding properties of the C-terminal PHD domains of the ING proteins are known, no structural information is available for the N-terminal domains. This domain contains a putative oligomerization site rich in helical structure in the ING2–5 members of the family. The N-terminal domain of ING4 was overexpressed in Escherichia coli and purified to homogeneity. Crystallization experiments yielded crystals that were suitable for high-resolution X-ray diffraction analysis. The crystals belonged to the orthorhombic space group C222, with unit-cell parameters a = 129.7, b = 188.3, c = 62.7 A. The self-rotation function and the Matthews coefficient suggested the presence of three protein dimers per asymmetric unit. The crystals diffracted to a resolution of 2.3 A using synchrotron radiation at the Swiss Light Source (SLS) and the European Synchrotron Radiation Facility (ESRF).

  • molecular basis of histone h3k4me3 recognition by ING4
    Journal of Biological Chemistry, 2008
    Co-Authors: Alicia Palacios, Ines G Munoz, Guillermo Montoya, David Pantojauceda, Maria J Marcaida, Daniel J Torres, Jose M Martingarcia, Irene Luque, Francisco J Blanco
    Abstract:

    The inhibitors of growth (ING) family of tumor suppressors consists of five homologous proteins involved in chromatin remodeling. They form part of different acetylation and deacetylation complexes and are thought to direct them to specific regions of the chromatin, through the recognition of H3K4me3 (trimethylated K4 in the histone 3 tail) by their conserved plant homeodomain (PHD). We have determined the crystal structure of ING4-PHD bound to H3K4me3, which reveals a tight complex stabilized by numerous interactions. NMR shows that there is a reduction in the backbone mobility on the regions of the PHD that participate in the peptide binding, and binding affinities differ depending on histone tail lengths Thermodynamic analysis reveals that the discrimination in favor of methylated lysine is entropy-driven, contrary to what has been described for chromodomains. The molecular basis of H3K4me3 recognition by ING4 differs from that of ING2, which is consistent with their different affinities for methylated histone tails. These differences suggest a distinct role in transcriptional regulation for these two ING family members because of the antagonistic effect of the complexes that they recruit onto chromatin. Our results illustrate the versatility of PHD fingers as readers of the histone code.

  • solution structure and nmr characterization of the binding to methylated histone tails of the plant homeodomain finger of the tumour suppressor ING4
    FEBS Letters, 2006
    Co-Authors: Alicia Palacios, Pascal Garcia, Daniel Padro, Eva Lopezhernandez, Irene Martin, Francisco J Blanco
    Abstract:

    Plant homeodomain (PHD) fingers are frequently present in proteins involved in chromatin remodelling, and some of them bind to histones. The family of proteins inhibitors of growth (ING) contains a PHD finger that bind to histone-3 trimethylated at lysine 4, and those of ING1 and ING2 also act as nuclear phosphoinositide receptors. We have determined the structure of ING4 PHD, and characterised its binding to phosphoinositides and histone methylated tails. In contrast to ING2, ING4 is not a phosphoinositide receptor and binds with similar affinity to the different methylation states of histone-3 at lysine 4.