Accumulation of cytolytic CD8+ T cells in B16-melanoma and proliferation of mature T cells in TIS21-knockout mice after T cell receptor stimulation.

In vivo and in vitro effects of TIS21 gene on the mature T cell activation and antitumor activities were explored by employing MO5 melanoma orthograft and splenocytes isolated from the TIS21-knockout (KO)(2) mice. Proliferation and survival of mature T cells were significantly increased in the KO than the wild type (WT3)e cells, indicating that TIS21 inhibits the rate of mature T cell proliferation and its survival. In MO5 melanoma orthograft model, the KO mice recruited much more CD8(+) T cells into the tumors at around day 14 after tumor cell injection along with reduced tumor volumes compared with the WT. The increased frequency of granzyme B+ CD8+ T cells in splenocytes of the KO mice compared with the WT may account for antitumor-immunity of TIS21 gene in the melanoma orthograft. In contrast, reduced frequencies of CD107a+ CD8+ T cells in the splenocytes of KO mice may affect the loss of CD8+ T cell infiltration in the orthograft at around day 19. These results indicate that TIS21 exhibits antiproliferative and proapoptotic effects in mature T cells, and differentially affects the frequencies of granzyme B+ CD8+ T-cells and CD107a+ CD8+ T-cells, thus transiently regulating in vivo anti-tumor immunity.

C-terminus-deleted FoxM1 is expressed in cancer cell lines and induces chromosome instability.

Forkhead Box M1 (FoxM1) protein is a transcription factor and regulates cell cycle. It is commonly upregulated in human cancer tissue and correlated with poor prognosis, suggesting that the overexpression of FoxM1 plays a critical role in carcinogenesis. In this study, we report the identification and characterization of a new variant of FoxM1, which was first isolated from our laboratory in hepatoma cell lines. Compared with wild-type FoxM1, the new variant lacks of C-terminus of FoxM1 (FoxM1ΔC), which is a transactivation domain. Reverse transcription-polymerase chain reaction and western blot analysis demonstrated that FoxM1ΔC was highly expressed in a variety of cancer cell lines such as HepG2, HeLa, A549, MB231, EJ, U2OS, Hep3B and MCF7, but not expressed in normal human dermal fibroblast (HDF). Immunoprecipitation assay revealed that FoxM1ΔC interacted with wild-type FoxM1. Furthermore, FoxM1ΔC bound to FoxM1 targeted gene promoter region and correlated with dysregulation of wild-type FoxM1. FoxM1ΔC delayed G2/M to G1 progression of cell cycle, decreased Aurora B(T232) phosphorylation and increased chromosome centromere interspace. Finally, FoxM1ΔC induced instability of chromosome and formation of aneuploid cells within 1 month when expressed in HDF. In conclusion, FoxM1ΔC is expressed in cancer cells and dysregulates normal cell cycle and induces chromosome instability.

Signals regulating necrosis of cardiomyoblast by BTG2(/TIS21/PC3) via activation of GSK3ß and opening of mitochondrial permeability transition pore in response to H2O2.

To investigate signal transduction pathway of cell death regulated by a tumor suppressor after oxidative stress, cardiomyoblasts were virally transfected with BTG2(/TIS21/PC3) (BTG2) and subsequently treated with H2O2. Heart muscle rarely expresses BTG2 unless oxidative stress occurs, however, ischemia induced BTG2 expression and necrosis, not apoptosis, of cardiomyoblasts. BTG2-expressioning cardiomyblasts showed impaired recoveries of survival kinases, Akt and Erk, thus sustaining GSK-3ß activity in 30 min of H2O2 exposure, in contrast to their rapid recoveries in LacZ control. The phenomenon was accompanied by the failure of ATP regeneration and the sustained activation of AMPK in the BTG2 expresser. Furthermore, H2O2 treatment markedly induced BTG2 translocation from nuclei to mitochondria along with cell death by cyclophilin D activation and mPTP opening. Exogenous and endogenous effect of BTG2 was confirmed by chemical inhibitors and BTG2-KO-MEF, respectively. Here, we suggest tumor suppressor, BTG2, as one of the regulators of necrosis in myocardium via inhibiting Akt/Erk, but activating GSK3ß and cyclophilin D, which resulted in mPTP opening in response to H2O2.

B-cell translocation gene 2 mediates crosstalk between PI3K/Akt1 and NFκB pathways which enhances transcription of MnSOD by accelerating IκBα degradation in normal and cancer cells.

BACKGROUND: B-cell translocation gene 2 (BTG2) belongs to antiproliferative (ARPO) gene family and the expression of BTG2, human ortholog of rat PC3 and mouse TIS21 gene, has been shown to render cancer cells more sensitive to doxorubicin treatment by upregulating MnSOD expression without regulating any other reactive oxygen species (ROS) scavenging enzymes. RESULTS: In the present study, by employing exogenous and endogenous BTG2/TIS21/Pc3 expression by transfection and transduction analyses, and by knockdown of gene expression using RNA interference or using gene knockout cells, we observed that BTG2 increased the binding of activated NF-κB (p65/RelA) to the enhancer element of MnSOD gene in the 2nd intron, which was regulated by p-Akt1, and the induction of MnSOD by BTG2 was accompanied with subsequent downregulation of ROS level and cyclin B1 biosynthesis along with the increase of p21WAF1, resulting in the G2/M arrest independent of p53. CONCLUSIONS: These results show for the first time that BTG2 mediates crosstalk between PI3K-Akt1 and NF-κB pathways, which regulates p53-independent induction of G2/M phase arrest both in normal and cancer cells.

Proper regulation of ß-adrenergic signal requires Btg2 gene for lipolysis and thermogenesis in response to starvation or cold acclimation in female mice.

Mammals maintain constant body temperature in cold environment by activating thermogenesis via adrenergic/protein kinase A (PKA) signaling. B-cell translocation gene 2 (BTG2/Tis21), induced by PKA signaling, regulates glucose and lipid metabolism in liver, yet its role in lipolysis and in thermogenesis is not explored. Here, Btg2-knockout (KO) mice failed to maintain body temperature under starvation, or in cold acclimation. And norepinephrine-induced thermogenic response was turned off earlier in the KO mice. Gender specifically, gonadal white adipose tissues (gWAT) of female-KO were very active in lipolysis in fed state, however, the fat degradation was diminished upon fasting or cold acclimation. Also, insulin sensitivity was increased in female-KO, but not in male-KO mice, along with the low bone mineral density and small brown adipose tissues (BAT). In the mechanistic aspect, expressions of UCP1 and lipases (LPL, ATGL, HSL) in gWAT of female-KO mice were significantly reduced in response to adrenergic signals. Here, we present some data that Btg2 gene is essential for properly respond to ß-adrenergic signals, and plays as a negative regulator of insulin signaling in female mice.

What matters in aging is signaling for responsiveness.

Biological responsiveness refers to the capacity of living organisms to adapt to changes in both their internal and external environments through physiological and behavioral mechanisms. One of the prominent aspects of aging is the decline in this responsiveness, which can lead to a deterioration in the processes required for maintenance, survival, and growth. The vital link between physiological responsiveness and the essential life processes lies within the signaling systems. To devise effective strategies for controlling the aging process, a comprehensive reevaluation of this connecting loop is imperative. This review aims to explore the impact of aging on signaling systems responsible for responsiveness and introduce a novel perspective on intervening in the aging process by restoring the compromised responsiveness. These innovative mechanistic approaches for modulating altered responsiveness hold the potential to illuminate the development of action plans aimed at controlling the aging process and treating age-related disorders.

Reduction of exportin 6 activity leads to actin accumulation via failure of RanGTP restoration and NTF2 sequestration in the nuclei of senescent cells.

We have previously reported that G-actin accumulation in nuclei is a universal phenomenon of cellular senescence. By employing primary culture of human diploid fibroblast (HDF) and stress-induced premature senescence (SIPS), we explored whether the failure of actin export to cytoplasm is responsible for actin accumulation in nuclei of senescent cells. Expression of exportin 6 (Exp6) and small G-protein, Ran, was significantly reduced in the replicative senescence, but not yet in SIPS, whereas nuclear import of actin by cofilin was already increased in SIPS. After treatment of young HDF cells with H(2)O(2), rapid reduction of nuclear RanGTP was observed along with cytoplasmic increase of RanGDP. Furthermore, significantly reduced interaction of Exp6 with RanGTP was found by GST-Exp6 pull-down analysis. Failure of RanGTP restoration was accompanied with inhibition of ATP synthesis and NTF2 sequestration in the nuclei along with accordant change of senescence morphology. Indeed, knockdown of Exp6 expression significantly increased actin molecule in the nuclei of young HDF cells. Therefore, actin accumulation in nuclei of senescent cells is most likely due to the failure of RanGTP restoration with ATP deficiency and NTF2 accumulation in nuclei, which result in the decrease of actin export via Exp6 inactivation, in addition to actin import by cofilin activation.

Loss of p21(Sdi1) expression in senescent cells after DNA damage accompanied with increase of miR-93 expression and reduced p53 interaction with p21(Sdi1) gene promoter.

To answer what is a critical event for higher incidence of tumor development in old than young individuals, primary culture of human diploid fibroblasts were employed and DNA damage was induced by doxorubicin or X-ray irradiation. Response to the damage was different between young and old cells; loss of p21(sdi1) expression in spite of p53(S¹5) activation in old cells along with [³H]thymidine and BrdU incorporation, but not in young cells. The phenomenon was confirmed by other tissue fibroblasts obtained from different donor ages. Induction of miR-93 expression and reduced p53 binding to p21 gene promoter account for loss of p21(sdi1) expression in senescent cells after DNA damage, suggesting a mechanism of in vivo carcinogenesis in aged tissue without repair arrest.

Role of TNF-α-Inducing Protein Secreted by Helicobacter pylori as a Tumor Promoter in Gastric Cancer and Emerging Preventive Strategies.

The tumor necrosis factor-α (TNF-α)-inducing protein (tipα) gene family, comprising Helicobacter pylori membrane protein 1 (hp-mp1) and tipα, has been identified as a tumor promoter, contributing to H. pylori carcinogenicity. Tipα is a unique H. pylori protein with no similarity to other pathogenicity factors, CagA, VacA, and urease. American H. pylori strains cause human gastric cancer, whereas African strains cause gastritis. The presence of Tipα in American and Euro-Asian strains suggests its involvement in human gastric cancer development. Tipα secreted from H. pylori stimulates gastric cancer development by inducing TNF-α, an endogenous tumor promoter, through its interaction with nucleolin, a Tipα receptor. This review covers the following topics: tumor-promoting activity of the Tipα family members HP-MP1 and Tipα, the mechanism underlying this activity of Tipα via binding to the cell-surface receptor, nucleolin, the crystal structure of rdel-Tipα and N-terminal truncated rTipα, inhibition of Tipα-associated gastric carcinogenesis by tumor suppressor B-cell translocation gene 2 (BTG2/TIS21), and new strategies to prevent and treat gastric cancer. Thus, Tipα contributes to the carcinogenicity of H. pylori by a mechanism that differs from those of CagA and VacA.

Skp2 enhances polyubiquitination and degradation of TIS21/BTG2/PC3, tumor suppressor protein, at the downstream of FoxM1.

TIS21(/BTG2/PC3) has been shown to work as a pan-cell cycle inhibitor and a negative regulator of cyclin B1/cdk1 and forkhead box M1 (FoxM1). Moreover, loss of TIS21 expression has been suggested as an early event in carcinogenesis of thymus, prostate, kidney, and liver. However, there is no report yet what regulates the in vivo stability of TIS21 protein. Here, TIS21 was found to be a target of ubiquitin ligase, S phase kinase associated protein 2 (Skp2), the expression of which was regulated by FoxM1. Leucine rich repeat (LRR) domain of Skp2 could bind to TIS21 C-terminus and facilitated TIS21 degradation via ubiquitin-proteasome pathway. Skp2 without LRR and C-terminus deleted TIS21 (TIS21DeltaC) failed to interact with each other, and failure of their interaction prolonged half-life of TIS21 protein. Furthermore, in vivo function of TIS21, inhibition of cell growth, was regulated by expressions of Skp2 and FoxM1; It was significantly enhanced by knock down of Skp2 expression in the TIS21 adenovirus infected cells, whereas it was significantly ameliorated by co-expression of FoxM1 with TIS21. These data indicate that TIS21 is a novel target of SCF-Skp2 ubiquitin ligase, which is regulated by expression of FoxM1.

Akt Downregulates B-Cell Translocation Gene-2 Expression Via Erk1/2 Inhibition for Proliferation of Cancer Cells.

B-cell translocation gene 2 (Btg2) is a tumor suppressor gene that is implicated in many biological processes. Akt is a serine/threonine kinase which was originally discovered as an oncogene. The prognostic value of Akt activation in some types of cancers and its effect on tumor suppressor genes remains to be fully elucidated. In the current research we have investigated the Akt-mediated downregulation of Btg2 that increased cells proliferation and cells survival. Human leukemia HL-60, THP-1 and colon cancer DLD-1 cells were used in this study. Inhibition of Akt with LY294002 significantly increased Btg2 mRNA expression while activation of Akt with insulin decreased Btg2 expression. Contrary to this, treatment of cells with U0126, a MAPK kinase inhibitor, significantly abrogated Btg2 expression. Moreover, LY294002 treatment increased Erk1/2 activation, decreased cells proliferation and cells viability while activation of Akt by insulin led to an increase in cells survival and cells division. Exogenous expression of Btg2 decreased cells proliferation both in the presence and absence of insulin and arrested cells at G1 phase. Akt negatively regulates Btg2 via Erk1/2 inhibition that lead to an increase in cells survival and cells proliferation. This elucidates a new mechanism for Btg2 regulation and Akt mediated tumorgenicity.

Mitochondrial nucleoid remodeling and biogenesis are regulated by the p53-p21WAF1-PKCζ pathway in p16INK4a-silenced cells.

Mitochondrial dysfunction is linked to age-related senescence phenotypes. We report here the pathway increasing nucleoid remodeling and biogenesis in mitochondria during the senescence of foreskin human diploid fibroblasts (fs-HDF) and WI-38 cells. Replicative senescence in fs-HDF cells increased mitochondrial nucleoid remodeling as indicated by 5-bromo-2'-deoxyuridine (BrdU) incorporation and mitochondrial transcription factor A (TFAM) expression in enlarged and fused mitochondria. Mitochondrial nucleoid remodeling was accompanied by mitochondrial biogenesis in old cells, and the expression levels of OXPHOS complex-I, -IV and -V subunits, PGC-1α and NRF1 were greatly increased compared to young cells. Activated protein kinase C zeta (PKCζ) increased mitochondrial activity and expressed phenotypes of delayed senescence in fs-HDF cells, but not in WI-38 cells. The findings were reproduced in the doxorubicin-induced senescence of young fs-HDF and WI-38 cells via the PKCζ-LKB1-AMPK signaling pathway, which was regulated by the p53-p21WAF1 pathway when p16INK4a was silenced. The signaling enhanced PGC-1α-NRF1-TFAM axis in mitochondria, which was demonstrated by Ingenuity Pathway Analysis of young and old fs-HDF cells. Activation of the p53-p21WAF1 pathway and silencing of p16INK4a are responsible for mitochondrial reprogramming in senescent cells, which may be a compensatory mechanism to promote cell survival under senescence stress.

TIS21 negatively regulates hepatocarcinogenesis by disruption of cyclin B1-Forkhead box M1 regulation loop.

UNLABELLED: A functional and biochemical interaction of TIS21(/BTG2/PC3) with Forkhead box M1 (FoxM1), essential transcription factor for hepatocyte regeneration and a master regulator of mitotic gene expression, was explored. Growth of hepatocellular carcinoma (HCC), developed by a single injection of diethylnitrosamine (DEN), was the same in both the TIS21(+/+) and TIS21(-/-) mice until 6 months, whereas it was significantly higher in the TIS21(-/-) mice at 9 months. Expression of TIS21 was significantly lower in both human and murine HCCs than in the surrounding tissues. Forced expression of TIS21 impaired growth, proliferation, and tumorigenic potential of Huh7 cells. At the mechanistic level, TIS21 inhibited FoxM1 phosphorylation, a required modification for its activation, by reducing cyclin B1-cdk1 activity, examined by in vitro kinase assay and FoxM1 mutant analyses. These observations were further confirmed in vivo by the reciprocal control of TIS21 expression and FoxM1 phosphorylation in the diethylnitrosamine-induced HCCs and TIS21(-/-) mouse embryonic fibroblast (MEF), in addition to increased expression of cyclin B1 and cdk1 activity. CONCLUSION: TIS21 negatively regulated hepatocarcinogenesis in part by disruption of the FoxM1-cyclin B1 regulatory loop, thereby inhibiting proliferation of transformed cells developed in mouse and human livers.

TIS21/(BTG2) negatively regulates estradiol-stimulated expansion of hematopoietic stem cells by derepressing Akt phosphorylation and inhibiting mTOR signal transduction.

It has been known that 12-O-tetradecanoyl phorbol-13-acetate-inducible sequence 21 (TIS21), ortholog of human B-cell translocation gene 2, regulates expansions of stage-specific thymocytes and hematopoietic progenitors. In the present study, lineage-negative (Lin(-))/stem cell antigen-1-positive (Sca-1+)/c-Kit+ (LSK) cell content was significantly elevated in bone marrow (BM) of TIS21-knockout (TIS21(-/-)) female mice, suggesting 17beta-estradiol (E(2))-regulated progenitor expansion. E(2) induced DNA synthesis and cell proliferation of mouse embryonic fibroblasts (MEFs) isolated from TIS21(-/-) mice, but not wild type (WT). In contrast to WT, E(2) failed to activate protein kinase B (Akt) in the TIS21(-/-) MEFs, independent of extracellular signal-regulated kinase 1/2 (Erk1/2) activation. Despite attenuation of Akt activation, mammalian target of rapamycin (mTOR) was constitutively activated in the TIS21(-/-) MEFs. Furthermore, mitogen-activated protein kinase 1/2 inhibitor or knockdown of Erk1 could restore activation of Akt and downregulate mTOR. Immunoprecipitation showed Akt preferentially bound to phosphorylated Erk1/2 (p-Erk1/2) in TIS21(-/-) cells, but reconstitution of TIS21 inhibited their interaction. E(2)-injected TIS21(-/-) male mice also increased LSK cells in BM. Taken together, expansion of hematopoietic progenitors in TIS21(-/-) female mice might be through inhibition of Akt activation, and constitutive activation of mTOR via preferential binding of TIS21 to E(2)-induced p-Erk1/2, compared with that of Akt. Our results suggest that TIS21 plays a pivotal role in maintaining the hematopoietic stem cell compartment and hematopoiesis.

Translational downregulation of Twist1 expression by antiproliferative gene, B-cell translocation gene 2, in the triple negative breast cancer cells.

Twist1, a key transcription factor regulating epithelial-mesenchymal transition and cancer metastasis, is highly expressed in invasive cancers in contrast to the loss of BTG2/TIS21 expression. Based on our observation that forced expression of BTG2/TIS21 downregulated Twist1 protein expression without altering mRNA level, we investigated molecular mechanisms of the BTG2/TIS21-inhibited Twist1 translation in the triple negative breast cancer (TNBC) cells and in vivo BTG2/TIS21-knockout (KO) mice and human breast cancer tissues. (1) C-terminal domain of Twist1 and Box B of BTG2/TIS21 interacted with each other, which abrogated Twist1 activity. (2) BTG2/TIS21 inhibited translational initiation by depleting eIF4E availability via inhibiting 4EBP1 phosphorylation. (3) Expression of BTG2/TIS21 maintained p-eIF2α that downregulates initiation of protein translation, confirmed by eIF2α-AA mutant expression and BTG2/TIS21 knockdown in MEF cells. (4) cDNA microarray analysis revealed significantly higher expression of initiation factors-eIF2A, eIF3A, and eIF4G2-in the BTG2/TIS21-KO mouse than that in the wild type. (5) BTG2/TIS21-inhibited translation initiation lead to the collapse of polysome formation and the huge peak of 80s monomer in the BTG2/TIS21 expresser, but not in the control. (6) mRNAs and protein expressions of elongation factors were also downregulated by BTG2/TIS21 expression in TNBC cells, but much higher in both TIS21-KO mice and lymph node-positive human breast cancers. (7) BTG2/TIS21-mediated Twist1 loss was not due to the protein degradation by ubiquitination and autophagy activation. (8) Twist1 protein level was significantly higher in various organs of TIS21-KO mice compared with that in the control, indicating the in vivo role of BTG2/TIS21 gene in the regulation of Twist1 protein level. Altogether, the present study support our hypothesis that BTG2/TIS21 is a promising target to combat with metastatic cancers with high level of Twist1 without BTG2/TIS21 expression.

Pleiotrophin inhibits transforming growth factor beta1-induced apoptosis in hepatoma cell lines.

Pleiotrophin (PTN) is a hepatocyte growth factor and considered to play roles in liver fibrogenesis and hepatocarcinogenesis. In this study we examined the mechanism of the action of PTN in these pathological processes. First, we confirmed that hepatic stellate cells (HSCs) and Kupffer cells, and also later hepatocytes in hyperplastic nodules increased PTN mRNA expressions during carbon tetrachloride-induced liver fibrosis. Then, the relationship between PTN and transforming growth factor beta1 (TGFbeta1), a known potent pro-fibrogenetic cytokine, in carcinogenesis was investigated using hepatoma cell lines. Huh-7 human hepatoma cells weakly expressed PTN, but HepG2 human hepatoma cells and FaO rat hepatoma cells did not. Recombinant (r) TGFbeta1 induced the cultured Huh-7 cells to undergo apoptosis, which was inhibited by rPTN. Huh-7 cells became resistant to TGFbeta1-, but not mitomycin C-induced apoptosis when transfected with PTN gene, indicating the specificity of the PTN anti-apoptotic activity. Poly ADP ribose polymerase, procaspase-8 and procaspase-3 were not cleaved in the TGFbeta1-reluctant cells. The TGFbeta1-induced caspase-3 activation was also suppressed in Huh-7 and FaO cells both transduced with PTN gene-bearing adenoviruses. In summary, PTN was expressed in HSCs, Kupffer cells, and hepatocytes in fibrotic liver. We propose that PTN specifically antagonizes the TGFbeta1 activity during liver fibrosis.

PPARα-Target Gene Expression Requires TIS21/BTG2 Gene in Liver of the C57BL/6 Mice under Fasting Condition.

The TIS21/BTG2/PC3 gene belongs to the antiproliferative gene (APRO) family and exhibits tumor suppressive activity. However, here we report that TIS21 controls lipid metabolism, rather than cell proliferation, under fasting condition. Using microarray analysis, whole gene expression changes were investigated in liver of TIS21 knockout (TIS21-KO) mice after 20 h fasting and compared with wild type (WT). Peroxisome proliferator-activated receptor alpha (PPARα) target gene expression was almost absent in contrast to increased lipid synthesis in the TIS21-KO mice compared to WT mice. Immunohistochemistry with hematoxylin and eosin staining revealed that lipid deposition was focal in the TIS21-KO liver as opposed to the diffuse and homogeneous pattern in the WT liver after 24 h starvation. In addition, cathepsin E expression was over 10 times higher in the TIS21-KO liver than that in the WT, as opposed to the significant reduction of thioltransferase in both adult and fetal livers. At present, we cannot account for the role of cathepsin E. However, downregulation of glutaredoxin 2 thioltransferase expression might affect hypoxic damage in the TIS21-KO liver. We suggest that the TIS21/BTG2 gene might be essential to maintain energy metabolism and reducing power in the liver under fasting condition.

TIS21/BTG2 inhibits breast cancer growth and progression by differential regulation of mTORc1 and mTORc2-AKT1-NFAT1-PHLPP2 signaling axis.

PURPOSE: It has been reported that PI3K/AKT pathway is altered in various cancers and AKT isoforms specifically regulate cell growth and metastasis of cancer cells; AKT1, but not AKT2, reduces invasion of cancer cells but maintains cancer growth. We propose here a novel mechanism of the tumor suppresser, TIS21/BTG2, that inhibits both growth and invasion of triple negative breast cancer cells via AKT1 activation by differential regulation of mTORc1 and mTORc2 activity. METHODS: Transduction of adenovirus carrying TIS21/BTG2 gene and transfection of short interfering RNAs were employed to regulate TIS21/BTG2 gene expression in various cell lines. Treatment of mTOR inhibitors and mTOR kinase assays can evaluate the role of mTORc in the regulation of AKT phosphorylation at S473 residue by TIS21/BTG2 in breast cancer cells. Open data and immunohistochemical analysis were performed to confirm the role of TIS21/BTG2 expression in various human breast cancer tissues. RESULTS: We observed that TIS21/BTG2 inhibited mTORc1 activity by reducing Raptor-mTOR interaction along with upregulation of tsc1 expression, which lead to significant reduction of p70S6K activation as opposed to AKT1S473, but not AKT2, phosphorylation via downregulating PHLPP2 (AKT1-specific phosphatase) in breast cancers. TIS21/BTG2-induced pAKTS473 required Rictor-bound mTOR kinase, indicating activation of mTORc2 by TIS21/BTG2 gene. Additionally, the TIS21/BTG2-induced pAKTS473 could reduce expression of NFAT1 (nuclear factor of activated T cells) and its target genes, which regulate cancer microenvironment. CONCLUSIONS: TIS21/BTG2 significantly lost in the infiltrating ductal carcinoma, but it can inhibit cancer growth via the TIS21/BTG2-tsc1/2-mTORc1-p70S6K axis and downregulate cancer progression via the TIS21/BTG2-mTORc2-AKT1-NFAT1-PHLPP2 pathway.

Inhibition of TNFα-interacting protein α (Tipα)-associated gastric carcinogenesis by BTG2/TIS21 via downregulating cytoplasmic nucleolin expression.

To understand the regulation of Helicobacter pylori (H. pylori)-associated gastric carcinogenesis, we examined the effect of B-cell translocation gene 2 (BTG2) expression on the biological activity of Tipα, an oncoprotein secreted from H. pylori. BTG2, the human ortholog of mouse TIS21 (BTG2/TIS21), has been reported to be a primary response gene that is transiently expressed in response to various stimulations. Here, we report that BTG2 is constitutively expressed in the mucous epithelium and parietal cells of the gastric gland in the stomach. Expression was increased in the mucous epithelium following H. pylori infection in contrast to its loss in human gastric adenocarcinoma. Indeed, adenoviral transduction of BTG2/TIS21 significantly inhibited Tipα activity in MKN-1 and MGT-40, human and mouse gastric cancer cells, respectively, thereby downregulating tumor necrosis factor-α (TNFα) expression and Erk1/2 phosphorylation by reducing expression of nucleolin, a Tipα receptor. Chromatin immunoprecipitation proved that BTG2/TIS21 inhibited Sp1 expression and its binding to the promoter of the nucleolin gene. In addition, BTG2/TIS21 expression significantly reduced membrane-localized nucleolin expression in cancer cells, and the loss of BTG2/TIS21 expression induced cytoplasmic nucleolin availability in gastric cancer tissues, as evidenced by immunoblotting and immunohistochemistry. Higher expression of BTG2 and lower expression of nucleolin were accompanied with better overall survival of poorly differentiated gastric cancer patients. This is the first report showing that BTG2/TIS21 inhibits nucleolin expression via Sp1 binding, which might be associated with the inhibition of H. pylori-induced carcinogenesis. We suggest that BTG2/TIS21 is a potential inhibitor of nucleolin in the cytoplasm, leading to inhibition of carcinogenesis after H. pylori infection.

Phylogenetic characterization of norovirus strains detected from sporadic gastroenteritis in Seoul during 2014-2016.

BACKGROUND: Phylogenetic analysis of norovirus (NoV) is efficient for tracking NoV transmission. To determine the widespread NoV strains in Seoul, we conducted an extensive phylogenetic characterization of NoV-positives from 1659 diarrheal specimens collected in 2014-2016 for the Seoul NoV-surveillance. RESULTS: When the large numbers of NoV partial VP1 genome sequences were analyzed in acute gastroenteritis patients along with the phylogenetic characterization, we could identify molecular epidemiologic patterns based on the genetic characteristics of sporadic NoV strains circulating in Seoul, which could provide a detailed description of the genome-wide and community-wide NoV evolution in each genotype. The average NoV detection rate in our study period was 16.34% that was increased by 7.44% from 13.17% in 2014 to 20.61% in 2016. Prevalence of NoV GI and GII was 4.43% and 93.36%, respectively, and the GII.4, GII.17, and GII.3 were found to be the major type among 17 genotypes of NoV. The most prevalent one was GII.4 (50.92%) that was followed by GII.17 (18.08%) and GII.3 (9.96%). According to an extensive phylogenetic analysis based on partial VP1 sequences of 1008 NoV (276 sporadic, 518 outbreak and 214 reference), pandemic strains of GII.17, GII.4 and GII.3 have emerged in succession during the 2014-2016 Seoul NoV-surveillance. GII.17 emerged as GII.17|Kawasaki323 in 2014, and became the predominant genotype in 2015 with GII.17|2014_Kawasaki lineages (CUHK-NS-616/Kawasaki308). The formerly predominant GII.4 remained high-level with GII.4|2012_Sydney in 2014 and internally replaced to GII.4|2016_Kawasaki194 lineage (NOR-2565/NOR-2558/OH16002) that caused the sporadic NoV explosion since December 2015. Sporadically prevalent GII.3|Hu/Aichio334-13/2013 failed to develop any outbreaks, whereas sporadic GII.3|Hu/3-28/2015/HNZZ/CHN caused heavy outbreaks in Seoul without preparation time since November 2016. CONCLUSIONS: This is the first extensive phylogenetic study revealing the important events of NoV strains circulating in Seoul. Particularly, our study period from 2014 to 2016 was very dynamic with the emergences of the three main NoV strains (GII.17|2014_Kawasaki, GII.4|2016_Kawasaki194 and GII.3|Hu/3-28/2015/HNZZ/CHN) every year. We are sure that it is hard to detect above findings by simple conventional analysis. Our present study reports a future paradigm of the NoV molecular epidemiology, which might be highly valuable to track new strains and predict oncoming outbreaks.